Scalp care solution with effects of controlling oil, relieving, inhibiting bacteria and regulating scalp micro-ecology as well as preparation method and application of scalp care solution

Abstract

The invention discloses a scalp care solution with the effects of controlling oil, relieving, inhibiting bacteria and adjusting scalp micro-ecology and a preparation method and application thereof.The preparation method of the scalp care solution comprises the following steps that a fermentation substrate is inoculated with saccharomycetes, primary fermentation culture is conducted for 24-72 h, then secondary fermentation culture is conducted through Lactobacillus gasseri, and the scalp care solution is obtained; wherein the fermentation substrate comprises rice, barley seeds and water; wherein the saccharomycetes comprise Saccharomyces cerevisiae (Saccharomyces cerevisiae) and Saccharomyces fibulligera (Saccharomyces fibulligera), and the saccharomycetes comprise Saccharomyces cerevisiae (Saccharomyces fibulligera) and Saccharomyces fibulligera (Saccharomyces fibulligera). The rice and the barley seeds are fermented in a microbial fermentation mode, the bioavailability of the rice and the barley seeds is improved, the content of active ingredients in the fermentation liquor prepared through the fermentation technology is increased, the fermentation liquor has the ideal effects of controlling oil, resisting inflammation, inhibiting bacteria, adjusting scalp micro-ecology and the like, the preparation technology is simple, and the method is suitable for industrial production. The fermentation conditions are mild, no organic solvent is used, and safety is achieved.

Description

Technical Field

[0001] This application belongs to the field of cosmetic technology, and in particular relates to a scalp care liquid with oil-controlling, soothing, antibacterial and scalp microecological regulation effects, as well as its preparation method and application. Background Technology

[0002] Modern lifestyles exacerbate problems such as scalp sensitivity, oiliness, and hair loss. Urban pollution, ultraviolet radiation, staying up late, and high stress lead to damage to the scalp barrier and bacterial imbalance, resulting in a high incidence of sensitivity, itching, and seborrheic dermatitis. Shampoo, as a cleanser with added nutrients, needs to be rinsed thoroughly with water after use. Due to its limited functionality, its conditioning effect is relatively low and it is difficult to meet the needs of hair root care.

[0003] Scalp care solution is a liquid product focused on scalp health. Its function is to cleanse the scalp while conditioning the scalp and hair roots, helping to prevent hair loss and reduce dandruff. Unlike shampoo, scalp care solution can be sprayed directly onto the hair roots without rinsing. Traditional scalp care solutions often contain extracts of Chinese herbs such as Polygonum multiflorum, Panax notoginseng, and Artemisia argyi to relieve itching and control oil. However, the large molecules in these extracts are relatively difficult to penetrate the scalp, resulting in low utilization and limited effectiveness.

[0004] Therefore, in order to fill the market gap and accurately solve the sub-health of the scalp, it is urgent to develop a scalp care solution that is both safe and easy to use, and has excellent effects in controlling oil and inhibiting bacteria and regulating the scalp microecology. Summary of the Invention

[0005] The technical problem this application aims to solve is to overcome the limitations of existing shampoos in terms of function, and the low bioavailability of traditional Chinese medicine compound scalp care solutions, which cannot be directly absorbed by the scalp and thus have limited effects. This application provides a scalp care solution with oil-controlling, soothing, antibacterial, and scalp microecological regulating effects, along with its preparation method and application. This application utilizes a secondary microbial fermentation process to ferment rice and barley seeds, improving their bioavailability. The fermented liquid obtained through this process has an increased content of active ingredients, exhibiting ideal oil-controlling, anti-inflammatory, antibacterial, and scalp microecological regulating effects. Furthermore, the preparation process is simple, the fermentation conditions are mild, and no organic solvents are used, reducing resource waste and achieving energy conservation and environmental protection.

[0006] This application adopts the following technical solution to solve the above-mentioned technical problems: This application provides a method for preparing a scalp care solution with oil-controlling, soothing, antibacterial, and scalp microecological regulation effects, comprising the following steps: inoculating yeast into a fermentation substrate, culturing it in a single fermentation culture for 24-72 hours, and then subjecting it to Lactobacillus gasseri (… Lactobacillus gasseriThe yeast was prepared by secondary fermentation; wherein the fermentation substrate included rice, barley seeds, and water; and the yeast included *Saccharomyces cerevisiae*. Saccharomyces cerevisiae ) and cladomycin ( Saccharomycopsis fibuligera ).

[0007] In some embodiments, the weight ratio of rice, barley seeds, and water in the fermentation substrate may be (0.33~10):(0.33~10):100, preferably (0.66~5):(0.66~5):100, for example 1:1:100.

[0008] In some embodiments, the rice is rice powder that has been pulverized and sieved; further, the particle size of the rice powder after sieving is 50 to 150 mesh.

[0009] In some embodiments, the barley seeds are crushed and sieved barley seed powder, and further, the particle size of the barley seed powder sieved is 50-150 mesh.

[0010] In some embodiments, the water may include purified water and / or deionized water.

[0011] In some embodiments, the method for preparing the fermentation substrate includes: mixing the components of the fermentation substrate evenly.

[0012] The mixing can be carried out under stirring conditions as is customary in the art, and the stirring speed can be 200~400 rpm, preferably 250~350 rpm, for example 300 rpm.

[0013] In some embodiments, the fermentation substrate further includes a sterilization process before use.

[0014] The sterilization method for the fermentation substrate is generally high-temperature sterilization.

[0015] When the fermentation substrate is sterilized using the high-temperature sterilization method, the sterilization temperature can be a temperature conventional for this type of operation in the art, preferably 110℃~135℃, more preferably 115℃~130℃, for example 121℃; the sterilization pressure can be a pressure conventional for this type of operation in the art, preferably 0.1~0.15MPa, more preferably 0.1~0.13MPa, for example 0.12MPa; and the sterilization time can be a time conventional for this type of operation in the art, preferably 20~45min, more preferably 25~40min, for example 30min.

[0016] In some embodiments, the ratio of viable cells of the Saccharomyces cerevisiae to the Saccharomyces cerevisiae is 1:(0.1~3), preferably 1:(0.5~2).

[0017] In some embodiments, the Saccharomyces cerevisiae includes Saccharomyces cerevisiae deposited at the China General Microbiological Culture Collection Center with accession number CGMCC NO. 17452.

[0018] The *Saccharomyces cerevisiae* strain with accession number CGMCC No. 17452 was accessed on March 27, 2019, and is classified as *Saccharomyces cerevisiae* (…). Saccharomyces cerevisiae The depository is China General Microbiological Culture Collection Center (CGMCC), located at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, 100101, China.

[0019] In some embodiments, the cladostomata-coated yeast includes the cladostomata-coated yeast deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 22671.

[0020] The *Saccharomyces cladoides* strain with accession number CGMCC No. 22671 was deposited on June 7, 2021, and is classified as *Saccharomyces cladoides*. Saccharomycopsis fibuligera The depository is named China General Microbiological Culture Collection Center (CGMCC), located at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, 100101, China.

[0021] In some embodiments, the yeast may be added in the form of a yeast culture, as is customary in the art.

[0022] The brewer's yeast is added in the form of a brewer's yeast liquid. The viable concentration of the brewer's yeast in the brewer's yeast liquid can be 10. 6 ~10 10 CFU / mL, preferably 10 7 ~10 9 CFU / mL.

[0023] In some embodiments, the cladosporium-coated yeast is added in the form of a cladosporium-coated yeast solution, wherein the viable concentration of the cladosporium-coated yeast in the cladosporium-coated yeast solution is 10. 5 ~10 10 CFU / mL, preferably 10 6 ~10 9 CFU / mL.

[0024] In some embodiments, the inoculation amount of the *Saccharomyces cerevisiae* broth, by volume percentage, is 0.2% to 5% (v / v), preferably 0.5% to 2% (v / v); the inoculation amount of the *Saccharomyces cerevisiae* broth, by volume percentage, is 0.2% to 5% (v / v), preferably 0.5% to 2% (v / v). In this invention, the inoculation amount (v / v) refers to the volume of *Saccharomyces cerevisiae* broth or *Saccharomyces cerevisiae* broth inoculated onto 100 mL of the fermentation substrate. For example, an inoculation amount of 0.5% (v / v) means that 0.5 mL of broth is inoculated into 100 mL of fermentation substrate.

[0025] In some embodiments, the yeast strain further includes a strain activation step before use. The strain activation can be performed using conventional activation methods in the art.

[0026] In some embodiments, the preparation method of the yeast culture can be conventional in the art and generally includes the following steps: inoculating the yeast into YPD solid medium for streak activation, culturing in an incubator at 28°C for 48 hours to obtain single colonies; picking the obtained single colonies with an inoculation loop into YPD liquid medium, culturing in a shaking incubator at 28°C and a rotation speed of 180 r / min for 36-72 hours, more preferably 48 hours, to obtain the yeast culture.

[0027] In some embodiments, the conditions and methods for the primary fermentation culture can be conventional in the art, and generally can be shaker culture.

[0028] When the primary fermentation culture is carried out using the shaker culture, the rotation speed of the shaker is 150~200 rpm, for example 180 rpm.

[0029] In some embodiments, the temperature of the primary fermentation culture can be 24~37°C, preferably 24~30°C.

[0030] In some embodiments, the primary fermentation culture time is preferably 24-48 hours.

[0031] In some embodiments, the Lactobacillus gasseri is inoculated into the fermentation broth obtained after the first fermentation is completed.

[0032] In some embodiments, the *Lactobacillus gasseri* includes *Lactobacillus gasseri* deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 34898. Lactobacillus gasseri ).

[0033] The Lactobacillus gasseri with accession number CGMCC No. 34898 was deposited on June 16, 2025, and is classified as follows: Lactobacillus gasseriThe depository is China General Microbiological Culture Collection Center (CGMCC), located at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, 100101, China.

[0034] In some embodiments, the Lactobacillus gasseri may be added in the form of Lactobacillus gasseri broth, as is customary in the art.

[0035] The viable bacterial concentration of *Lactobacillus gasseri* in the *Lactobacillus gasseri* culture may be 10. 5 ~10 10 CFU / mL, preferably 10 6 ~10 10 CFU / mL, more preferably 10 8 ~10 10 CFU / mL.

[0036] In some embodiments, the preparation method of the Lactobacillus gasseri bacterial solution can be conventional in the art and generally includes the following steps: dissolving the Lactobacillus gasseri powder in sterile deionized water.

[0037] In some embodiments, the inoculation amount of *Lactobacillus gasseri* culture is 0.2% to 5% (v / v), preferably 0.5% to 2% (v / v), by volume percentage. In this invention, the inoculation amount (v / v) refers to the volume of *Lactobacillus gasseri* culture added based on the fermentation broth obtained after the first fermentation. For example, an inoculation amount of 1.0% (v / v) means that 1.0 mL of *Lactobacillus gasseri* culture is added to 100 mL of the fermentation broth obtained after the first fermentation.

[0038] In some embodiments, before inoculating the *Lactobacillus gasseri*, the fermentation broth obtained after the first fermentation is further subjected to sterilization and cooling. The sterilization method can be a conventional high-temperature sterilization method used in the art. The cooling can be a conventional cooling to room temperature method used in the art.

[0039] Furthermore, when the fermentation broth obtained after the first fermentation is sterilized using the high-temperature sterilization method, the sterilization temperature can be a temperature conventional for this type of operation in the art, preferably 110℃~125℃, more preferably 115℃~121℃, for example 121℃; the sterilization time can be a time conventional for this type of operation in the art, preferably 15~35min, more preferably 15~25min, for example 20min.

[0040] In some embodiments, the secondary fermentation culture is a static culture.

[0041] In some embodiments, the temperature of the secondary fermentation culture can be 37~45°C, preferably 37~43°C, for example 40°C.

[0042] In some embodiments, the secondary fermentation culture time can be 12-24 hours, preferably 12-15 hours.

[0043] In some embodiments, the secondary fermentation process further includes sterilization, and the sterilization method can be a high-temperature sterilization method commonly used in the art.

[0044] When the high-temperature sterilization method is used to sterilize the secondary fermentation, the sterilization temperature can be the temperature conventional for this type of operation in the art, preferably 110℃~125℃, more preferably 115℃~121℃, for example 121℃, and the sterilization time can be the time conventional for this type of operation in the art, preferably 15~35min, more preferably 15~25min, for example 20min.

[0045] In some embodiments, after the sterilization operation following the secondary fermentation, the process may further include at least one of the following operations: cooling, separating and collecting the supernatant, and mixing with a preservative.

[0046] In this context, according to conventional practice, the cooling may be cooling to room temperature.

[0047] The separation can be performed using conventional separation methods in the art, such as centrifugation. The centrifugation speed can be a conventional speed for this type of operation in the art, preferably 3000~9000 rpm, more preferably 4000~6000 rpm, for example 4800 rpm; the centrifugation time can be a conventional time for this type of operation in the art, preferably 10~40 min, more preferably 20~40 min, for example 30 min.

[0048] Preferably, the centrifugation process may further include a re-sterilization operation, wherein the re-sterilization method is a conventional high-temperature sterilization method used in the art. When the re-sterilization is performed using the high-temperature sterilization method, the re-sterilization temperature is a temperature conventional for this type of operation in the art, preferably 110℃~125℃, more preferably 115℃~121℃, for example 121℃, and the re-sterilization time is a time conventional for this type of operation in the art, preferably 20~40 min, more preferably 25~35 min, for example 20 min.

[0049] During the mixing process with the preservative, the mixing temperature can be a temperature commonly used in this type of operation in the art, preferably 50~80℃, more preferably 70~80℃.

[0050] In the process of mixing with the preservative, the preservative may include p-hydroxyacetophenone and / or 1,2-hydroxyacetophenone, as is commonly known in the art. Hexanediol.

[0051] When the preservative includes p-hydroxyacetophenone, the mass percentage of p-hydroxyacetophenone in the supernatant of the sterilized material can be 0.5% to 1%.

[0052] When the preservative includes the 1, 2 In the case of hexanediol, the 1,2 The mass fraction of hexanediol in the supernatant of the sterilized material is 0.5% to 1%. In some embodiments, the room temperature generally refers to 15~40°C.

[0053] This application also provides a scalp care solution with oil-controlling, soothing, antibacterial, and scalp microecological regulation effects, which is prepared by the scalp care solution with oil-controlling, soothing, antibacterial, and scalp microecological regulation effects as described above.

[0054] This application also provides the use of the scalp care liquid described above, which has the effects of oil control, soothing, antibacterial and regulating scalp microecology, as a product, as an additive or as a base in the preparation of topical skin agents.

[0055] In some embodiments, the scalp care solution with oil-controlling and antibacterial effects may be at least one of the antibacterial active ingredient, oil-controlling active ingredient, anti-inflammatory active ingredient, and skin microecological regulating ingredient in the topical skin agent.

[0056] The antibacterial active ingredient may be an antibacterial active ingredient that inhibits Staphylococcus aureus and / or Malassezia.

[0057] The oil-controlling active ingredient may be an oil-controlling active ingredient that inhibits the proliferation of sebaceous gland cells.

[0058] The anti-inflammatory active ingredient may be an anti-inflammatory active ingredient that can clear the inflammatory factor IL-8.

[0059] This application also provides a topical skin agent comprising a scalp care solution as described above, which has the effects of oil control, soothing, antibacterial properties, and regulation of scalp microecology.

[0060] In some embodiments, the scalp care solution having oil-controlling, soothing, antibacterial, and scalp microecological regulating effects accounts for 5% to 100% of the mass percentage of the topical skin agent, preferably 5% to 90%.

[0061] In some embodiments, the scalp care solution with oil-controlling, soothing, antibacterial, and scalp microecological regulating effects may also include at least one of anti-allergic or soothing active ingredients, skin barrier repair active ingredients, and anti-acne active ingredients.

[0062] In some embodiments, the scalp care solution with oil-controlling, soothing, antibacterial, and scalp microecological regulating effects can be shampoo, conditioner, hair mask, serum, spray, or scalp care nutrient solution, etc.

[0063] Based on common knowledge in the field, the above-mentioned preferred conditions can be combined arbitrarily to obtain various preferred embodiments of this application.

[0064] All reagents and raw materials used in this application are commercially available.

[0065] The positive and progressive effects of this application are as follows: (1) This application first uses Saccharomyces cerevisiae and Saccharomyces cerevisiae to ferment rice and barley seeds, and then uses Lactobacillus gasseri for secondary fermentation. This can decompose the plant cell wall and fully release the active ingredients in rice and barley seeds. This not only reduces the loss of active ingredients, but also makes the fermentation product have excellent antibacterial, oil-controlling, anti-inflammatory and scalp microecological regulation effects. It can significantly reduce the survival rate of sebaceous gland cells and inhibit the expression of Staphylococcus aureus, Malassezia and inflammatory factor IL-8. The final product can be used directly as a scalp care solution or added to the basic formula of cosmetics.

[0066] (2) This application utilizes microorganisms to ferment rice and barley seeds. No chemical components are added during the preparation of fermentation product filtrate. The fermentation product filtrate prepared has good safety and skin feel. Moreover, the fermentation conditions are mild, the fermentation cycle is short, and the production cost is low, which is conducive to large-scale production. As a scalp care liquid, it expands the application field of rice and barley seeds. Attached Figure Description

[0067] This application can be better understood by referring to the description given below in conjunction with the accompanying drawings. These drawings, together with the detailed description below, are included in and form part of this specification, and are used to further illustrate preferred embodiments of the application and explain the principles and advantages of the application. Wherein: Figure 1 Comparative graphs showing the inhibitory effects of the scalp care solutions prepared in Examples 1-3 and Comparative Examples 1-6 on Staphylococcus aureus; Figure 2 Comparison of the oil-controlling effects of the scalp care solutions prepared in Examples 1-3 and Comparative Examples 1-6; Figure 3 Comparative diagrams showing the inhibition of inflammatory factor IL-8 expression by the scalp care solutions prepared in Examples 1-3 and Comparative Examples 1-6; Figure 4 The graph shows a comparison of the inhibitory effects of the scalp care solutions prepared in Examples 1-3 and Comparative Examples 1-6 on Malassezia. Detailed Implementation

[0068] The present application is further illustrated below by way of embodiments, but these embodiments are not intended to limit the scope of the present application. Experimental methods not specifically described in the following embodiments are performed according to conventional methods and conditions, or as selected in accordance with the product instructions.

[0069] The raw materials used in the following examples and comparative examples are all commercially available.

[0070] In the following examples, Saccharomyces cerevisiae ( Saccharomyces cerevisiae The deposit date of the sample was March 27, 2019, the accession number was CGMCC No. 17452, and the classification name was *Saccharomyces cerevisiae* (Saccharomyces cerevisiae). Saccharomyces cerevisiae The depository is China General Microbiological Culture Collection Center (CGMCC), located at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, 100101, China.

[0071] In the following examples, the *Saccharomyces cladoides* strain was deposited on June 7, 2021, with accession number CGMCC No. 22671, and classified as *Saccharomyces cladoides* (…). Saccharomycopsis fibuligera The depository is named China General Microbiological Culture Collection Center (CGMCC), located at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, 100101, China.

[0072] In the following examples, Lactobacillus gasseri ( Lactobacillus gasseri The deposit date is June 16, 2025, the accession number is CGMCC No. 34898, and the classification name is... Lactobacillus gasseri The depository is China General Microbiological Culture Collection Center (CGMCC), located at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, 100101, China.

[0073] In the following examples, *Lactobacillus plantarum* ( Lactiplantibacillus plantarum The deposit date of the specimen is October 27, 2023, the accession number is CGMCC No.28799, the depositary is the China General Microbiological Culture Collection Center (CGMCC), the address is No.3, No.1 Beichen West Road, Chaoyang District, Beijing, 100101.

[0074] In the following examples, Saccharomyces cerevisiae ( Saccharomyces cerevisiaePreparation of bacterial culture: Saccharomyces cerevisiae was inoculated into YPD solid medium for streak activation and cultured at 28℃ for 48 hours to obtain single colonies. Single colonies were then picked up using an inoculation loop and placed into YPD liquid medium, and cultured at 28℃ and 180 rpm for 48 hours in a shaking incubator, yielding approximately 10 viable cells. 8 Saccharomyces cerevisiae culture at CFU / mL.

[0075] In the following examples, the preparation of *Saccharomyces cerevisiae* culture medium was as follows: *Saccharomyces cerevisiae* was inoculated into YPD solid medium for streak activation and cultured at 28°C for 48 hours to obtain single colonies; the obtained single colonies were picked up with an inoculation loop and placed into YPD liquid medium, and cultured in a shaking incubator at 28°C and 180 r / min for 48 hours to obtain approximately 10 viable cells. 8 CFU / mL of capsule-coated yeast culture.

[0076] In the following examples, the preparation of Lactobacillus gasseri bacterial suspension was as follows: Lactobacillus gasseri powder was dissolved in sterile water to obtain a viable count of approximately 10. 9 CFU / mL Lactobacillus gasseri culture.

[0077] In the following examples, the preparation of *Lactobacillus plantarum* bacterial suspension was carried out by dissolving *Lactobacillus plantarum* powder in sterile water to obtain a viable count of approximately 10. 9 CFU / mL of Lactobacillus plantarum bacterial solution.

[0078] Example 1 (1) First, the rice and barley seeds were crushed and passed through a 100-mesh sieve. 300 mL of deionized water, 3 g of rice powder, and 3 g of barley seed powder were mixed evenly. After mixing evenly, the mixture was sterilized in a high-pressure autoclave at 121℃ for 20 min. After sterilization, it was cooled to room temperature to obtain the initial fermentation system. Saccharomyces cerevisiae and Saccharomyces thuringiensis were inoculated into the initial fermentation system for fermentation. The viable cell count of both Saccharomyces cerevisiae and Saccharomyces thuringiensis was 10. 8 The bacterial culture was added at a concentration of CFU / mL, with a volume of 1.5 mL. After inoculation, the bacterial culture was shaken well and fermented in a constant temperature incubator at 28℃ for 48 h at a shaking speed of 180 r / min. (2) After fermentation, sterilize in a high-pressure steam sterilizer at 121°C for 20 minutes. After sterilization, cool to room temperature and then add 3 mL of live bacteria at a concentration of 10. 9CFU / mL of Lactobacillus gasseri culture was shaken well and subjected to secondary fermentation at 40℃ for 15 hours. After fermentation, the culture was sterilized at 121℃ for 20 minutes. After sterilization, the culture was cooled to room temperature and centrifuged at 4800 rpm for 30 minutes. After centrifugation, the supernatant was collected and sterilized again at 121℃ for 30 minutes. Based on the mass of the collected supernatant, 0.5% p-hydroxyacetophenone and 0.5% 1,2-hexanediol were added and mixed at 75℃. The mixture was thoroughly mixed to obtain a scalp care solution with oil-controlling, soothing, antibacterial, and scalp microecological regulating effects.

[0079] Example 2 Compared with Example 1, the only difference is that 1 mL of Saccharomyces cerevisiae culture and 2 mL of saccharomyces cerevisiae culture are added during the fermentation process in step (1), and other conditions and parameters are the same as in Example 1.

[0080] Example 3 Compared with Example 1, the only difference is that 2 mL of Saccharomyces cerevisiae culture and 1 mL of saccharomyces cerevisiae culture are added during the fermentation process in step (1), and other conditions and parameters are the same as in Example 1.

[0081] Comparative Example 1 Compared with Example 1, the only difference is that fermentation is not performed; all other conditions and parameters are the same as in Example 1. The specific operation is as follows: First, grind the rice and barley seeds separately through a 100-mesh sieve. Take 300mL of deionized water, 3g of rice powder, and 3g of barley seed powder, and mix them evenly. After mixing evenly, sterilize in an autoclave at 121℃ for 20min. After sterilization, cool to room temperature, centrifuge to collect the supernatant at 4800r / min for 30min. After centrifugation, sterilize again at 121℃ for 30min. After sterilization, based on the mass of the collected supernatant, add 0.5% p-hydroxyacetophenone and 0.5% 1,2-hexanediol, and mix at 75℃. Mix evenly.

[0082] Comparative Example 2 Compared to Example 1, the only difference is that a secondary fermentation process is not performed (i.e., no inoculation with Lactobacillus gasseri for secondary fermentation); all other conditions and parameters are the same as in Example 1. The specific operation is as follows: First, rice and barley seeds were separately pulverized and passed through a 100-mesh sieve. 300 mL of deionized water, 3 g of rice powder, and 3 g of barley seed powder were mixed thoroughly. The mixture was then sterilized in a 121℃ autoclave for 20 minutes. After sterilization, it was cooled to room temperature to obtain the initial fermentation system. *Saccharomyces cerevisiae* and *Saccharomyces cerevisiae* were inoculated into the initial fermentation system for fermentation. The viable cell count of both the *Saccharomyces cerevisiae* and *Saccharomyces cerevisiae* cultures was 10⁻⁶. 8 The bacterial culture was added at a concentration of CFU / mL, with each addition being 1.5 mL. After inoculation, the mixture was shaken well and fermented in a constant temperature incubator at 28℃ for 48 h at a shaking speed of 180 r / min. After fermentation, the mixture was sterilized at 121℃ for 20 min. After sterilization, the mixture was cooled to room temperature and centrifuged at 4800 r / min for 30 min. After centrifugation, the supernatant was collected and sterilized again at 121℃ for 30 min. After sterilization, based on the mass of the collected supernatant, 0.5% p-hydroxyacetophenone and 0.5% 1,2-hexanediol were added and mixed at 75℃.

[0083] Comparative Example 3 Compared to Example 1, the only difference is that a primary fermentation process is omitted (i.e., fermentation is not carried out by inoculating with *Saccharomyces cerevisiae* and *Saccharomyces cerevisiae*, but directly by inoculating with *Lactobacillus gasseri*). All other conditions and parameters are the same as in Example 1. The specific operation is as follows: First, the rice and barley seeds were separately ground and passed through a 100-mesh sieve. 300 mL of deionized water, 3 g of rice powder, and 3 g of barley seed powder were mixed thoroughly. The mixture was then sterilized in a 121℃ autoclave for 20 minutes. After sterilization, it was cooled to room temperature to obtain the initial fermentation system. 3 mL of live bacteria at a concentration of 10⁻⁶ were then added to the initial fermentation system. 9 CFU / mL of Lactobacillus gasseri culture was shaken well and fermented. The fermentation conditions were: static fermentation at 40℃ for 15 hours. After fermentation, the culture was sterilized at 121℃ for 20 minutes. After sterilization, the culture was cooled to room temperature and centrifuged at 4800 rpm for 30 minutes. After centrifugation, the supernatant was collected and sterilized again at 121℃ for 30 minutes. After sterilization, based on the mass of the collected supernatant, 0.5% p-hydroxyacetophenone and 0.5% 1,2-hexanediol were added and mixed at 75℃. The mixture was then thoroughly mixed.

[0084] Comparative Example 4 Compared with Example 1, the only difference is that only 6g of rice powder is used as the fermentation substrate, while other conditions and parameters are the same as in Example 1.

[0085] Comparative Example 5 Compared with Example 1, the only difference is that Lactobacillus gasseri is replaced with Lactobacillus plantarum CGMCC NO.28799, and the other conditions and parameters are the same as in Example 1.

[0086] Comparative Example 6 Compared with Example 1, the difference is that in the secondary fermentation process, the fermentation is carried out in a shaker at a speed of 180 rpm, and other conditions and parameters are the same as in Example 1.

[0087] Example 1: Antibacterial efficacy test (inhibition of Staphylococcus aureus experiment) The inhibition rate of the samples prepared in Examples 1-3 and Comparative Examples 1-6 against Staphylococcus aureus was determined to evaluate their antibacterial activity.

[0088] Experimental materials Staphylococcus aureus strain, liquid culture medium, test samples, 96-well plate, sterile physiological saline, sterile pipettes and tips, incubator, microplate shaker, microplate reader Experimental steps 1. Bacterial culture: The cryopreserved Staphylococcus aureus strain was inoculated into the culture medium and cultured at 37°C with shaking for 24 hours.

[0089] 2. Bacterial suspension dilution: Dilute to a McFarland value of 0.5. A McFarland value of 0.5 is 10. 8 CFU / mL, i.e., OD 600 It is between 0.08 and 0.13.

[0090] 3. Sample addition: Add the sample to the corresponding well of the 96-well plate. (Sample + bacterial suspension) Add bacterial suspension to each well to achieve a final bacterial concentration of approximately 10. 6 CFU / mL. - Fill the blank control wells with sterile saline (saline + bacterial suspension).

[0091] 4. Incubation: Place the 96-well plate in a 37°C incubator for 24 hours.

[0092] 5. After incubation, the OD values ​​of the wells at 600 nm were measured using a microplate reader. The results are shown in Table 1 and [Table data missing]. Figure 1 .

[0093] Antibacterial rate = (OD blank control group - OD sample group) / OD blank control group × 100%.

[0094] Table 1

[0095] From Table 1 and Figure 1The results showed that the Staphylococcus aureus inhibition rates in Examples 1-3 were significantly higher than those in Comparative Examples 1-6. This indicates that the Staphylococcus aureus inhibition effects of the examples were superior to those of Comparative Examples 1-6, demonstrating that the scalp care solution prepared in this application has excellent Staphylococcus aureus inhibition effects. Example 2: Oil control efficacy test (inhibition of sebaceous gland cell proliferation) Cell viability (inhibition of sebaceous gland cell proliferation) This experiment used SZ95 cells from the Chinese Scientific Cell Bank to verify the cell viability of the products prepared in Examples 1-3 and Comparative Examples 1-6.

[0096] Experimental steps: The products obtained in Examples 1-3 and Comparative Examples 1-6 were respectively prepared into 1% (v / v) experimental test solutions using serum-free DMEM medium. Human sebaceous gland cells were cultured in a medium containing 10% fetal bovine serum and 1% penicillin-antibody (1×10⁻⁶) 5 Cells were cultured in DMEM medium containing 100 mg / L penicillin and 100 mg / L streptomycin. Cells were grown in a 37°C, 5% CO2 saturated humidity incubator. When cell confluence reached 85% or higher, logarithmic growth phase cells were digested with 0.05% trypsin, and the digestion was terminated with serum-containing DMEM. Cells were counted using a cell counting chamber, and the cell suspension concentration was adjusted to 8 × 10⁻⁶ cells / mL. 4 Cells were seeded at a rate of 100 μL / well in 96-well plates and incubated at 37°C and 5% CO2 for 12 h. The old culture medium was removed, and the cells were washed twice with phosphate-buffered saline (PBS). 100 μL of the filtered and sterilized test solution was added to each well of the experimental group, with three replicates per test solution. The control group contained cells and was inoculated with serum-free DMEM medium. The blank control group contained no cells and was inoculated with 100 μL of PBS. The cells were then incubated at 37°C and 5% CO2 for 24 h. Then, 10 μL of CCK-8 solution was added to each well, and the cells were incubated for another 3 h. The absorbance was measured at 450 nm, and the cell viability was calculated. The results are shown in Table 2. Figure 2 .

[0097] The formula for calculating cell viability is as follows: Cell viability (%) = (A experimental group - A blank control group) / (A control group - A blank control group) × 100%.

[0098] Table 2

[0099] From Table 2 and Figure 2The results showed that the sebaceous gland cell survival rate of Examples 1-3 was significantly lower than that of Comparative Examples 1-6, indicating that the scalp care solution prepared in Examples 1-3 has a significant scalp oil control effect.

[0100] Example 3: Anti-inflammatory effect (inhibition of IL-8 expression) Log-phase HaCaT cells were seeded at a density of 250,000 cells / mL into 6-well cell culture plates, with 2 mL of cell suspension added to each well. The plates were cultured for 12 hours. For the model group and sample group, 2 mL of 10... 6 Cells were incubated with a CFU / mL Staphylococcus aureus suspension for 24 hours. In the control group, 2 mL of serum-free DMEM culture medium was added and the cells were incubated for 24 hours. The bacterial suspension was discarded and the cells were washed twice with PBS. In the model group, 2 mL of serum-free DPEM culture medium was added and the cells were incubated for 24 hours. In the control group, 2 mL of serum-free DMEM culture medium was added and the cells were incubated for 24 hours. In the sample group, the sample diluted with serum-free DMEM culture medium (sample volume fraction of 1%) was added and the cells were incubated for 24 hours.

[0101] Preparation of cell lysis supernatant: The IP lysis buffer was prepared at a ratio of IP:PMSF of 100:1. After cell culture was completed, the culture supernatant was collected, and the cells were washed 2-3 times with PBS pre-cooled to 4°C. The PBS was discarded, and 500 μL of lysis buffer was added to each well. After the cells were fully lysed, the cell lysis buffer and cell pellet were collected with a cell scraper. The cells were centrifuged at 12000 r / min for 20 min, and the supernatant was collected for later use.

[0102] The inflammatory factors were measured according to the ELISA kit instructions. OD values ​​were measured at 450 nm. Results are shown in Table 3. Figure 3 .

[0103] Table 3

[0104] From Table 3 and Figure 3 The results showed that the IL-8 content of the inflammatory factor in Examples 1-3 was significantly lower than that in Comparative Examples 1-6, indicating that the scalp care solution prepared in Examples 1-3 has a significant anti-inflammatory effect.

[0105] Example 4: Inhibition Efficacy Test (Inhibition of Malassezia) Malassezia is a lipophilic fungus belonging to the yeast family. It relies on lipids in sebum as a nutrient source for growth and thrives in oily environments. In most cases, Malassezia lives in a symbiotic relationship with its host. However, when the host's immunity is weakened or sebum secretion is excessive, Malassezia proliferates abnormally, leading to the accumulation of its breakdown products, unsaturated fatty acids (such as oleic acid). These fatty acids irritate the scalp, causing inflammation and accelerating the shedding of scalp keratinocytes, which can even lead to hair loss in severe cases.

[0106] To determine the inhibition rate of the sample against Malassezia in order to evaluate its antibacterial activity.

[0107] Experimental materials Dixon broth base (liquid medium for culturing Malassezia), modified Dixon agar (MD) (solid medium, inverted plates, smeared), Tween 40 (added to the solid medium), and PBS. Experimental procedure. Experimental steps: (1) Prepare the liquid and solid culture media required for Malassezia according to the instructions for use, and use them after sterilization; (2) Resuscitate the Malassezia strain, add it to the liquid culture medium, and carry out scale-up culture at 37°C and 120 r. (3) The scalp care solutions prepared in Examples 1-3 and Comparative Examples 1-6 were filtered through a 0.22 μM organic phase microporous membrane, and 100 μL of the bacterial suspension cultured overnight (1×10⁻⁶) was taken. 7 The CFU / mL concentration was evenly spread on an agar plate; (4) Take 11 sterile filter paper discs with a diameter of 6 mm and place them on the spread agar plate. Add 20 μL of PBS and ketoconazole (0.1 mg / ml) to the filter paper for the blank group and positive control group, respectively. Add 20 μL of sample to the filter paper for the sample group. (5) After incubating it in a 37℃ incubator for 48 hours, measure the diameter of its inhibition zone and calculate its inhibition rate. Each group of experiments was repeated 3 times.

[0108] (6) Calculate the inhibition rate based on the diameter of the inhibition zone: Inhibition rate (%) = (Dt) Dc) / Dc×100, Dt: diameter of inhibition zone in experimental group (mm), Dc: diameter of inhibition zone in control group (mm), results are shown in Table 4 and Figure 4 .

[0109] Table 4

[0110] From Table 4 and Figure 4The results showed that the Malassezia inhibition rate of Examples 1-3 was significantly higher than that of Comparative Examples 1-6, indicating that the scalp care solution prepared in Examples 1-3 could significantly inhibit the proliferation of Malassezia.

[0111] Finally, it should be noted that the terms “comprising,” “including,” or any other variations thereof in this application are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such process, method, article, or apparatus. Although this application has been disclosed above through the description of specific embodiments, it should be understood that those skilled in the art can devise various modifications, improvements, or equivalents to this application within the spirit and scope of the appended solutions. Such modifications, improvements, or equivalents should also be considered to be included within the scope of protection claimed in this application.

Claims

1. A method for preparing a scalp care solution with oil-controlling, soothing, antibacterial, and scalp microecological regulating effects, comprising the following steps: inoculating yeast into a fermentation substrate, culturing it for 24-72 hours in a single fermentation, and then subjecting it to Lactobacillus gasseri (… Lactobacillus gasseri It was prepared by secondary fermentation culture; among which, The fermentation substrate includes rice, barley seeds, and water; the yeast includes Saccharomyces cerevisiae (Saccharomyces cerevisiae). Saccharomyces cerevisiae ) and cladomycin ( Saccharomycopsis fibuligera ).

2. The method for preparing the scalp care solution according to claim 1, characterized in that, The weight ratio of rice, barley seeds, and water in the fermentation substrate is (0.33~10):(0.33~10):100, preferably (0.66~5):(0.66~5):100; And / or, the rice is rice powder that has been crushed and sieved; further, the particle size of the rice powder after sieving is 50 to 150 mesh. And / or, the barley seeds are barley seed powder that has been crushed and sieved, and further, the particle size of the barley seed powder sieved is 50 to 150 mesh; And / or, the water includes purified water and / or deionized water; And / or, the fermentation substrate may also include a sterilization process before use.

3. The method for preparing the scalp care solution according to claim 1, characterized in that, The live cell ratio of the *Saccharomyces cerevisiae* and *Saccharomyces cerevisiae* is 1:(0.1~3), preferably 1:(0.5~2). And / or, the Saccharomyces cerevisiae includes the Saccharomyces cerevisiae deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC NO. 17452; And / or, the capsule-coated yeast includes the capsule-coated yeast deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 22671; And / or, the brewer's yeast is added in the form of brewer's yeast liquid, and further, the viable concentration of the brewer's yeast in the brewer's yeast liquid is 10. 6 ~10 10 CFU / mL, preferably 10 7 ~10 9 CFU / mL; And / or, the *Saccharomyces cerevisiae* is added in the form of a *Saccharomyces cerevisiae* liquid, and further, the viable concentration of the *Saccharomyces cerevisiae* in the *Saccharomyces cerevisiae* liquid is 10. 5 ~10 10 CFU / mL, preferably 10 6 ~10 9 CFU / mL; And / or, by volume percentage, the inoculation amount of the Saccharomyces cerevisiae liquid is 0.2%~5% (v / v), preferably 0.5%~2% (v / v); the inoculation amount of the saccharomyces cerevisiae liquid is 0.2%~5% (v / v), preferably 0.5%~2% (v / v).

4. The method for preparing the scalp care solution according to any one of claims 1-3, characterized in that, The primary fermentation culture was a shaker culture. And / or, when the primary fermentation culture is carried out using the shaker culture, the rotation speed of the shaker is 150~200 rpm; And / or, the temperature of the primary fermentation culture is 24~37℃, preferably 24~30℃; And / or, the fermentation time for the first fermentation is 24-48 hours.

5. The method for preparing the scalp care solution according to any one of claims 1-4, characterized in that, The Lactobacillus gasseri was inoculated into the fermentation broth obtained after the first fermentation was completed; And / or, the Lactobacillus gasseri includes Lactobacillus gasseri deposited at the China General Microbiological Culture Collection Center with accession number CGMCC No. 34898; And / or, the Lactobacillus gasseri is added in the form of Lactobacillus gasseri bacterial suspension; And / or, the viable concentration of *Lactobacillus gasseri* in the *Lactobacillus gasseri* culture is 10. 5 ~10 10 CFU / mL, preferably 10 6 ~10 10 CFU / mL, preferably 10 8 ~10 10 CFU / mL; And / or, by volume percentage, the inoculation amount of the Lactobacillus gasseri culture is 0.2% to 5% (v / v), preferably 0.5% to 2% (v / v). And / or, before inoculating the lactobacillus, the fermentation broth obtained after the first fermentation is sterilized and cooled; And / or, the secondary fermentation culture is a static culture; And / or, the temperature of the secondary fermentation culture is 37~45℃, preferably 37~43℃, for example 40℃; And / or, the secondary fermentation culture time is 12~24h, preferably 12~15h.

6. The method for preparing the scalp care solution according to any one of claims 1-5, characterized in that, The secondary fermentation process also includes sterilization, and the sterilization method is high-temperature sterilization. Further, when the high-temperature sterilization method is used for sterilization after the secondary fermentation, the sterilization temperature is 110℃~125℃, more preferably 115℃~121℃; and the sterilization time is 15~35min, more preferably 15~25min. And / or, after the sterilization operation following the secondary fermentation, the process further includes at least one of the following operations: cooling, separating and collecting the supernatant, and mixing with a preservative; And / or, the cooling may be cooling to room temperature; And / or, the separation is performed by centrifugation; further, the centrifugation speed is 3000~9000 rpm, more preferably 4000~6000 rpm; the centrifugation time is 10~40 min, more preferably 20~40 min; Preferably, the centrifugation process further includes a re-sterilization operation, wherein the re-sterilization method is high-temperature sterilization; further, when the high-temperature sterilization method is used for the re-sterilization, the re-sterilization temperature is 110℃~125℃, more preferably 115℃~121℃; and the re-sterilization time is 20~40min, more preferably 25~35min. And / or, during the mixing process with the preservative, the mixing temperature is 50~80°C, more preferably 70~80°C; And / or, the preservative includes p-hydroxyacetophenone and / or 1,2 Hexanediol; Further, the p-hydroxyacetophenone accounts for 0.5%~1% of the mass percentage of the supernatant obtained after sterilization; the 1,2 The mass fraction of hexanediol in the supernatant obtained after sterilization is 0.5% to 1%; And / or, the room temperature is 15~40℃.

7. A scalp care solution with oil-controlling, soothing, antibacterial, and scalp microecological regulating effects, prepared by the method of any one of claims 1-6.

8. The application of a scalp care liquid as described in claim 7, which has the effects of oil control, soothing, antibacterial properties and regulation of scalp microecology, as a product, as an additive, or as a base in the preparation of topical skin agents; Furthermore, the scalp care liquid with oil-controlling and antibacterial effects can be used as at least one of the antibacterial active ingredients, oil-controlling active ingredients, anti-inflammatory active ingredients, and skin microecological regulating ingredients in the topical skin agent; Furthermore, the antibacterial active ingredient may be an antibacterial active ingredient that inhibits Staphylococcus aureus and / or Malassezia. Furthermore, the oil-controlling active ingredient may be an oil-controlling active ingredient that inhibits the proliferation of sebaceous gland cells; Furthermore, the anti-inflammatory active ingredient may be an anti-inflammatory active ingredient that can clear the inflammatory factor IL-8.

9. A topical skin agent, characterized in that, This includes the scalp care solution described in claim 7, which has the effects of oil control, soothing, antibacterial properties, and regulation of scalp microecology.

10. The topical skin agent according to claim 9, characterized in that, The scalp care solution with oil-controlling, soothing, antibacterial, and scalp microecological regulating effects accounts for 5% to 100% of the mass of the topical skin agent, preferably 5% to 90%. And / or, the scalp care liquid with oil-controlling, soothing, antibacterial and scalp microecological regulation effects further includes at least one of anti-allergic or soothing active ingredients, skin barrier repair active ingredients, and anti-acne active ingredients; And / or, the scalp care liquid with oil-controlling, soothing, antibacterial and scalp microecological regulating effects is at least one of shampoo, conditioner, hair mask, serum, spray or scalp care nutrient solution.

Images