A plant-based oil control acne-fighting composition, and methods of making and using the same

Abstract

This invention belongs to the field of cosmetic technology, specifically relating to a plant-based oil-controlling and acne-removing composition, its preparation method, and its application. The plant-based oil-controlling and acne-removing composition is selected from at least five of the following: Artemisia argyi extract, Saururus chinensis extract, Magnolia officinalis leaf extract, Lysimachia christinae extract, Commiphora flavescens extract, Forsythia suspensa extract, Sophora flavescens extract, Salix babylonica bark extract, and Rhodiola rosea extract. The plant-based oil-controlling and acne-removing composition provided by this invention has significant oil-controlling and acne-removing effects.

Description

Technical Field

[0001] This invention belongs to the field of cosmetic technology, specifically relating to a plant-based oil-controlling and acne-removing composition, its preparation method, and its application. Background Technology

[0002] People with oily, sensitive, and acne-prone skin are often troubled by skin problems such as redness, inflammation, excessive sebum secretion, and acne / pimples. Traditional sensitive skin products mostly focus on barrier repair and cannot solve the problems of excessive sebum secretion and acne / pimples. Currently, oil-controlling and acne-reducing products on the market are mainly divided into two categories. One category uses chemically synthesized oil-controlling and acne-reducing ingredients, such as retinoid derivatives, benzoyl peroxide, or salicylic acid. Although these products can effectively control oil and reduce acne, they are highly irritating and often cause problems such as dryness, redness, peeling, and barrier damage in people with oily and acne-prone skin. The other category uses plant extracts. These products have received widespread attention from people with oily and acne-prone skin and researchers because of their gentleness.

[0003] Using a combination of multiple plant extracts to achieve multiple effects is a common practice in this field. However, the plant extracts in existing compound plant-based oil-controlling and acne-removing compositions often employ traditional plant extraction processes (water decoction, organic solvent extraction), which generally result in problems such as low content of active ingredients and poor permeability, leading to unsatisfactory oil-controlling and acne-removing effects.

[0004] Chinese Patent Publication No. CN 121570401 A discloses a herbal acne-clearing composition, its preparation method, and its application. The herbal acne-clearing composition comprises: 1-3 parts of honeysuckle flower extract, 4-6 parts of magnolia bark extract, 0.1-1 parts of dandelion extract, and 2-5 parts of fermentation extract. The fermentation strain used in the fermentation extract is *Lactobacillus fermentum*, with accession number CGMCC No. 21381. The fermentation substrate powder includes *Phellodendron amurense* bark powder and *Salix babylonica* bark powder. The herbal acne-clearing composition disclosed in this technical solution has significant acne-removing effects and is gentle and non-irritating. Chinese patent CN 121910644A discloses an acne-repairing composition based on a six-dimensional synergistic effect. Its active ingredients include 1 to 10 parts of sturgeon caviar extract, 0.1 to 10 parts of chlorogenic acid, 0.1 to 20 parts of 4-tert-butylcyclohexanol, and 0.1 to 20 parts of willow extract. The sturgeon caviar extract is obtained through supercritical CO2 fluid extraction, and the willow extract is treated with a combined enzymatic hydrolysis process to achieve a salicin content of not less than 1.5 wt%. At least one active ingredient in the composition undergoes supramolecular inclusion modification. The composition is prepared sequentially, then mixed sequentially, and finally subjected to an inclusion reaction. This composition achieves a six-dimensional synergistic effect through antibacterial, oil-controlling, anti-inflammatory, soothing, keratin-clearing, barrier-repairing, and skin-maintaining system, constructing a complete acne-repairing and skin health maintenance system. It features highly effective acne treatment, gentle safety, and a low recurrence rate, making it suitable for various skin types, including sensitive acne-prone skin, and can be widely applied to various acne-repairing product formulations.

[0005] Although the above technical solution uses a novel extraction method and has the effect of removing acne, the fermentation extraction method takes a long time and the supercritical CO2 fluid extraction is costly.

[0006] In view of this, the present invention is hereby proposed. Summary of the Invention

[0007] The purpose of this invention is to provide a plant-based oil-controlling and acne-removing composition, its preparation method, and its application. The plant-based oil-controlling and acne-removing composition provided by this invention is formulated with specific plant raw materials and extracted using a supramolecular solvent-complex enzyme preparation enzymatic hydrolysis-octanol-thymol solution containing phosphatidylcholine, which has significant oil-controlling and acne-removing effects.

[0008] To achieve the above objectives, the present invention provides the following technical solution: The first aspect of the present invention provides a plant-based oil-controlling and acne-removing composition, comprising, by weight percentage, 3-5% of a compound plant extract, 2-3% of Primula veris extract, 1-2% of Symphytum officinale extract, and solvent to make up to 100%.

[0009] As a preferred embodiment, the plant raw materials of the compound plant extract are composed of Artemisia septithrum, Saururus chinensis and Magnolia grandifola leaves in a mass ratio of 4-6:2-3:2-3.

[0010] As a preferred embodiment, the preparation method of the compound plant extract is as follows: S1. Crush and sieve the dried plant material to obtain raw material powder; S2. The raw material powder is extracted sequentially using supramolecular solvent, enzymatic hydrolysis with a compound enzyme preparation, and extraction with an octanol-thymol solution containing phosphatidylcholine. The extracts are combined, concentrated and dried under reduced pressure to obtain the compound plant extract.

[0011] As a preferred embodiment, the supramolecular solvent is an organic acid-hydroxytyrosol aqueous solvent.

[0012] As a preferred embodiment, the supramolecular solvent is prepared by mixing an organic acid, hydroxytyrosol, and water, heating to 50-60°C, and reacting under nitrogen protection for 2-4 hours.

[0013] As a preferred embodiment, the molar ratio of the organic acid to hydroxytyrosol is 1-3:1-3.

[0014] As a preferred embodiment, the supramolecular solvent has a water content of 10-20 wt%.

[0015] As a preferred embodiment, the organic acid is selected from at least one of citric acid, lactic acid, malic acid, and succinic acid.

[0016] As a preferred embodiment, the compound enzyme preparation is composed of cellulase and pectinase in a mass ratio of 1:0.3-0.5.

[0017] As a preferred embodiment, the method for preparing the octanol-thymol solution containing phosphatidylcholine is as follows: octanol and thymol are mixed and stirred at 70-80℃ for 2-4 hours, and then phosphatidylcholine is added and mixed evenly to obtain the solution.

[0018] As a preferred embodiment, the molar ratio of octanol to thymol is 1:2-4.

[0019] As a preferred embodiment, the octanol-thymol solution containing phosphatidylcholine contains 2-4 wt% phosphatidylcholine.

[0020] As a preferred embodiment, step S2 specifically involves: mixing the raw material powder and supramolecular solvent, ultrasonically extracting, adding a compound enzyme preparation, enzymatic hydrolysis, ultrasonically inactivating the enzyme, filtering, and obtaining a first extract and filter residue; mixing the filter residue with an octanol-thymol solution containing phosphatidylcholine, stirring and extracting, filtering, and obtaining a second extract; combining the first and second extracts, concentrating and drying under reduced pressure to obtain the compound plant extract.

[0021] As a preferred embodiment, the ratio of the raw material powder to the supramolecular solvent is 1g:10-20mL.

[0022] As a preferred embodiment, the ultrasonic extraction temperature is 30-45℃ and the time is 40-90 min.

[0023] As a preferred embodiment, the ultrasonic power of the ultrasonic extraction is 80-150W.

[0024] As a preferred embodiment, the mass ratio of the compound enzyme preparation to the raw material powder is 1-2:100.

[0025] As a preferred embodiment, the enzymatic hydrolysis temperature is 30-55℃ and the time is 30-60 min.

[0026] As a preferred embodiment, the ultrasonic enzyme inactivation power is 400-500W and the time is 5-10min.

[0027] As a preferred embodiment, the ratio of the filter residue to the octanol-thymol solution containing phosphatidylcholine is 1g:10-20mL.

[0028] As a preferred embodiment, the stirring extraction temperature is 50-65℃, the time is 30-60 min, and the stirring speed is 100-300 rpm.

[0029] As a preferred embodiment, the solvent is butanediol and water in a mass ratio of 20-40:60-80.

[0030] The second aspect of the present invention provides a method for preparing a plant-based oil-controlling and acne-removing composition, comprising the following steps: mixing a compound plant extract, a herb extract, a herb extract, and a solvent evenly to obtain the composition.

[0031] The third aspect of this invention provides the application of a plant-based oil-controlling and acne-removing composition in the preparation of oil-controlling and acne-removing products.

[0032] As a preferred option, the oil-controlling and acne-removing products include, but are not limited to, serums, lotions, creams, and masks.

[0033] As a preferred embodiment, the amount of the plant-based oil-controlling and acne-removing composition added to the oil-controlling and acne-removing product is 0.5-3.0%.

[0034] Compared with the prior art, the advantages and beneficial effects of the present invention are as follows: The components in the plant-based oil-controlling and acne-removing composition provided by this invention do not act independently or are simply a collection of ingredients, but rather have a comprehensive effect of synergistic oil control, antibacterial, anti-inflammatory, and repairing properties: 1. The raw materials in the compound plant extract of the present invention are Artemisia argyi, Saururus chinensis and Magnolia officinalis leaves. Experiments have shown that the three are indispensable, interact with each other, complement each other, and together achieve significant oil control and acne removal effects.

[0035] 2. The compound plant extract of the present invention employs a specific extraction process: sequential extraction with a supramolecular solvent, enzymatic hydrolysis with a compound enzyme preparation, and extraction with an octanol-thymol solution containing phosphatidylcholine. The supramolecular solvent is an organic acid-hydroxytyrosol aqueous solvent, which can not only improve the extraction rate of active substances by breaking adsorption bonds and solubilizing, but also specifically bind to the active substances in the compound plant, improving the purity and activity of the active substances. Enzymatic hydrolysis with the compound enzyme preparation provides favorable conditions for subsequent extraction with an octanol-thymol solution containing phosphatidylcholine. Further extraction with an octanol-thymol solution containing phosphatidylcholine utilizes the solubilizing effect of phosphatidylcholine and the synergistic effect of the octanol-thymol system to enhance the dissolution effect of lipid-soluble active substances not extracted by the supramolecular solvent, achieving full extraction and effective utilization of the effective substances in the plant raw materials.

[0036] 3. The plant-based oil-controlling and acne-removing composition of the present invention uses extracts of *Hedyotis diffusa*, *Comfrey*, and other compound plant extracts. Experiments have shown that compared with plant extracts with similar effects, the three have a better synergistic effect and together achieve a significant oil-controlling and acne-removing effect.

[0037] 4. The plant-based oil-controlling and acne-removing composition provided by the present invention has an inhibition rate of >33% on cell lipids at a concentration of 0.2%, an inhibition rate of >96% on Propionibacterium acnes at a concentration of 2%, and an inhibition rate of >55% on 5α-reductase. Attached Figure Description

[0038] Figure 1 VISIA images of some subjects at different time points for the essence water prepared using the plant-based oil-controlling and acne-removing composition of Example 3; Figure 2 VISIA images of some subjects at different time points for the essence water prepared using the plant-based oil-controlling and acne-removing composition of Example 3 – porphyrin. Detailed Implementation

[0039] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0040] To further illustrate the present invention, the following embodiments provide a detailed description. All raw materials used in the following embodiments of the present invention are commercially available products; the companies listed below represent one of the ways to purchase these raw materials.

[0041] Yellow-flowered nine-ring grass extract: extraction solvent: water-ethanol, product specification 10:1, Shaanxi Dongjiang Kangtai Health Industry Co., Ltd.

[0042] Polysaccharifolia extract: Extraction solvent: water-ethanol, product specification 10:1, Xi'an Changyue Biotechnology Co., Ltd.

[0043] Sophora flavescens extract, extraction solvent: water, product specification 10:1, Shaanxi Runze Xinqi Biotechnology Co., Ltd.

[0044] Rhodiola rosea extract, extraction solvent: water, product specification 10:1, Shaanxi Runze Xinqi Biotechnology Co., Ltd.

[0045] Forsythia suspensa extract, extraction solvent: water, product specification 10:1, Shaanxi Runze Xinqi Biotechnology Co., Ltd.

[0046] Salix Alba bark extract, solvent: water, product specification 10:1, Hunan Kangjiutang Biotechnology Co., Ltd.

[0047] Commercially available Artemisia argyi extract from Central Asia: Extraction solvent: water, product specification 10:1, Shaanxi Sinote Biotechnology Co., Ltd.

[0048] Commercially available Sanbaicao extract: Extraction solvent: water, product specification 10:1, Shaanxi Runze Xinqi Biotechnology Co., Ltd.

[0049] Commercially available lotus and magnolia leaf extract: extraction solvent: water, product specification 10:1, Shaanxi Sinote Biotechnology Co., Ltd.

[0050] Cellulase: Yisheng Biotechnology (Shanghai) Co., Ltd., Cellulase R-10.

[0051] Pectinase: Yisheng Biotechnology (Shanghai) Co., Ltd., Pectinase Y-23.

[0052] Preparation example: Compound Plant Extract I: The plant raw materials consist of Artemisia argyi, Saururus chinensis, and Magnolia officinalis leaves in a mass ratio of 5:3:1. The preparation method is as follows: S1. Crush and sieve the dried plant material to obtain raw material powder; S2. Mix the raw material powder and supramolecular solvent at a ratio of 1g:15mL, and extract by ultrasonication at 40℃ and 100W for 60min. Add the compound enzyme preparation (the mass ratio of compound enzyme preparation to raw material powder is 1.5:100), and enzymatically hydrolyze at 40℃ for 60min. Inactivate the enzyme by ultrasonication, filter, and obtain the first extract and filter residue. Mix the filter residue and octanol-thymol solution containing phosphatidylcholine at a ratio of 1g:15mL, and extract by stirring at 60℃ and 200rpm for 60min. Filter to obtain the second extract. Combine the first extract and the second extract, concentrate and dry under reduced pressure to obtain the compound plant extract.

[0053] The supramolecular solvent is prepared by mixing citric acid, hydroxytyrosol, and water, heating to 60°C, and reacting under nitrogen protection for 4 hours.

[0054] The molar ratio of citric acid to hydroxytyrosol is 1:1, and the water content of the supramolecular solvent is 15wt%.

[0055] The compound enzyme preparation consists of cellulase and pectinase in a mass ratio of 1:0.4.

[0056] The method for preparing the octanol-thymol solution containing phosphatidylcholine is as follows: octanol and thymol are mixed and stirred at 75°C for 3 hours, then phosphatidylcholine is added and mixed evenly to obtain the solution.

[0057] The molar ratio of octanol to thymol is 1:3.

[0058] The octanol-thymol solution containing phosphatidylcholine contains 3 wt% phosphatidylcholine.

[0059] Compound Plant Extract II: The only difference from Compound Plant Extract I is that the plant raw materials are composed of Artemisia argyi and Magnolia officinalis leaves in a mass ratio of 8:1; all other aspects are the same.

[0060] Compound Plant Extract III: The only difference from Compound Plant Extract I is that the plant raw materials are composed of Saussurea costatum and Magnolia officinalis leaves in a mass ratio of 3:6; all other aspects are the same.

[0061] Compound Plant Extract IV: The only difference from Compound Plant Extract I is that the plant raw materials are composed of Artemisia argyi and Saururus chinensis in a mass ratio of 5:4; otherwise, they are the same.

[0062] Compound Plant Extract V: The only difference between Compound Plant Extract I and Compound Plant Extract V is the preparation method: S1. Crush and sieve the dried plant material to obtain raw material powder; S2. Mix the raw material powder and supramolecular solvent at a ratio of 1g:15mL, and extract ultrasonically at 40℃ and 100W for 60min. Add the compound enzyme preparation (the mass ratio of compound enzyme preparation to raw material powder is 1.5:100), and enzymatically hydrolyze at 40℃ for 60min. Inactivate the enzyme with ultrasound, filter, and obtain the first extract and filter residue. Mix the filter residue and supramolecular solvent at a ratio of 1g:15mL, and extract by stirring at 60℃ and 200rpm for 60min. Filter to obtain the second extract. Combine the first and second extracts, concentrate and dry under reduced pressure to obtain the compound plant extract. The rest are the same.

[0063] Complex Plant Extract VI: The only difference from Complex Plant Extract I is that the preparation method is as follows: S1. Crush and sieve the dried plant material to obtain raw material powder; S2. Mix the raw material powder and the octanol-thymol solution containing phosphatidylcholine at a ratio of 1g:15mL, and extract by ultrasonication at 40℃ and 100W for 60min. Add the compound enzyme preparation (the mass ratio of the compound enzyme preparation to the raw material powder is 1.5:100), and enzymatically hydrolyze at 40℃ for 60min. Inactivate the enzyme by ultrasonication, filter, and obtain the first extract and filter residue. Mix the filter residue and the octanol-thymol solution containing phosphatidylcholine at a ratio of 1g:15mL, and extract by stirring at 60℃ and 200rpm for 60min. Filter to obtain the second extract. Combine the first and second extracts, concentrate and dry under reduced pressure to obtain the compound plant extract. The rest are the same.

[0064] Compound Plant Extract VII: The only difference from Compound Plant Extract I is that the supramolecular solvent is replaced with 1,3-propanediol in the preparation process; otherwise, they are the same.

[0065] Compound Plant Extract VIII: The only difference from Compound Plant Extract I is that the octanol-thymol solution containing phosphatidylcholine is replaced with an octanol-thymol solution; all other aspects are the same.

[0066] Compound Plant Extract IX: The only difference from Compound Plant Extract I is that it is composed of commercially available Artemisia argyi extract, Saururus chinensis extract and Magnolia officinalis leaf extract in a mass ratio of 5:3:1; all other aspects are the same.

[0067] Example 1 A plant-based oil-controlling and acne-removing composition, by weight percentage, comprises 3% of compound plant extract I, 3% of *Hedyotis diffusa* extract, 2% of *Comfrey sphaerocephala* extract, and solvent to make up the balance to 100%.

[0068] The solvent is butanediol and water in a mass ratio of 30:70.

[0069] The preparation method of the above-mentioned plant-based oil-controlling and acne-removing composition includes the following steps: mixing compound plant extract I, yellow clover extract, comfrey extract and solvent evenly to obtain the final product.

[0070] Example 2 The only difference from Example 1 is that, by mass percentage, it includes 4% of compound plant extract I, 2.5% of *Gynostemma pentaphyllum* extract, 1.5% of *Comfrey* extract, and solvent to make up to 100%; all other aspects are the same.

[0071] Example 3 The only difference from Example 1 is that, by mass percentage, it includes 5% of compound plant extract I, 2% of *Hedyotis diffusa* extract, 1% of *Comfrey* extract, and solvent to make up to 100%; all other components are the same.

[0072] Example 4 The only difference from Example 3 is that Compound Plant Extract I is replaced with Compound Plant Extract II; all else is the same.

[0073] Example 5 The only difference from Example 3 is that compound plant extract I is replaced with compound plant extract III; all else is the same.

[0074] Example 6 The only difference from Example 3 is that compound plant extract I is replaced with compound plant extract IV; all other aspects are the same.

[0075] Example 7 The only difference from Example 3 is that compound plant extract I is replaced with compound plant extract V; all else is the same.

[0076] Example 8 The only difference from Example 3 is that compound plant extract I is replaced with compound plant extract VI; all else is the same.

[0077] Example 9 The only difference from Example 3 is that compound plant extract I is replaced with compound plant extract VII; all else is the same.

[0078] Example 10 The only difference from Example 3 is that Compound Plant Extract I is replaced with Compound Plant Extract VIII; all else is the same.

[0079] Example 11 The only difference from Example 3 is that compound plant extract I is replaced with compound plant extract IX; all other aspects are the same.

[0080] Example 12 The only difference from Example 3 is that the extract of *Hedyotis diffusa* is replaced with extract of *Sophora flavescens*; all other aspects are the same.

[0081] Example 13 The only difference from Example 3 is that the comfrey extract is replaced with Rhodiola rosea extract; all other aspects are the same.

[0082] Example 14 The difference from Example 3 is that, by mass percentage, it includes 5% Forsythia suspensa extract, 5% Sophora flavescens extract, 5% Salix babylonica bark extract, 2% Rhodiola rosea extract, and solvent to make up to 100%.

[0083] The solvent is butanediol and water in a mass ratio of 30:70.

[0084] The preparation method of the above-mentioned plant-based oil-controlling and acne-removing composition includes the following steps: mixing Forsythia suspensa extract, Sophora flavescens extract, Salix babylonica bark extract, Rhodiola rosea extract and solvent evenly to obtain the composition.

[0085] Performance Test 1: Acne-Clearing Efficacy (Sebum Droplets) 1. Test Objective and Principle 1.1 Test Objective This test is based on the sodium palmitate-induced immortalized sebaceous gland cell (SZ95) model. By examining the effect of the sample on cell lipid production, the acne-removing efficacy of the test sample is evaluated.

[0086] 1.2 Test Principle Human skin sebum can be divided into sebaceous gland sebum and epidermal sebum based on its source. Sebaceous glands are the main source of sebum, and they are distributed throughout the body except for the palms and soles. There can be as many as 400-900 sebaceous glands per cm² on the human face. 2 Under normal circumstances, sebum combines with sweat to form a sebum film to maintain and prevent skin moisture loss and resist external invasion. The fatty acids in sebum can also resist the invasion of harmful bacteria. However, excessive sebum secretion and changes in sebum composition may induce skin microbiome dysbiosis. Sebum participates in the growth and proliferation of Propionibacterium acnes, biofilm formation, and inflammation, thereby causing acne. Increased levels of some lipid components can also aggravate acne development. Free fatty acids such as palmitic acid and oleic acid can also activate the AMPK / SREBP-1 / FAS pathway, promoting increased lipid synthesis and metabolism in sebaceous gland cells and producing inflammatory responses.

[0087] Nile red is a lipophilic fluorescent dye that exhibits strong orange-red fluorescence at an excitation wavelength of 530 nm after binding to lipids. Therefore, the skin acne-reducing efficacy of samples can be evaluated to some extent by detecting changes in the fluorescence intensity of Nile red-labeled lipids in human sebaceous gland cells. This study established a sodium palmitate-induced sebum secretion model in human immortalized sebaceous gland cells (SZ95) and assessed the acne-reducing efficacy of the test samples by examining their effect on cellular lipid production.

[0088] 2. Test materials 2.1 Test System The cells used in this test were immortalized human sebaceous gland cells (SZ95).

[0089] 2.2 Main Reagents DMEM culture medium, fetal bovine serum, trypsin-EDTA, sodium palmitate, and Nile red.

[0090] 2.3 Main Equipment CO2 incubator, biosafety cabinet, inverted microscope, multi-functional microplate reader, inverted fluorescence microscope.

[0091] 2.4 Solution Preparation Sample solution: Take the sample solution to be tested, filter it through a 0.22um filter membrane for sterilization, and dilute the culture medium to 0.2% (v / v); Sodium palmitate solution: Prepare a 6mM sodium palmitate solution. When using, dilute the culture medium to 180uM.

[0092] 3. Testing Methods 1) Sample processing and grouping: This experiment used a 0.2% concentration sample group, and set up a blank control group and a model control group (180uM sodium palmitate).

[0093] 2) Cell seeding: Take cells in the logarithmic growth phase, seed them in a 24-well plate, add 1 mL of cell suspension to each well, and incubate in an incubator (37℃, 5% CO2) until the cell confluence reaches 50%-60%.

[0094] 3) Cell drug delivery: Add sample solution containing sodium palmitate, and set up a blank control group. After drug delivery, incubate the 24-well plate in an incubator (37℃, 5% CO2) for 48 hours.

[0095] 4) Observation and recording: Discard the original culture medium, rinse each well twice with an appropriate amount of PBS, add 500uL of Nile Red working solution and incubate in an incubator (37℃, 5% CO2) for 15min, rinse each well three times with an appropriate amount of PBS, observe and photograph under a fluorescence microscope (objective lens 20x), and use software to perform statistical analysis on the fluorescence images.

[0096] 5) Data statistics and analysis: Relative fluorescence intensity value = Fluorescence intensity value of sample group (model control group) / Fluorescence intensity value of blank control group Cellular lipid inhibition rate = (relative fluorescence intensity value of model control group - relative fluorescence intensity value of sample group) / relative fluorescence intensity value of model control group × 100%.

[0097] 4. Test Results Table 1 Performance Test 2: Acne-Clearing Efficacy (Quantitative Antibacterial Test of Propionibacterium acnes Suspension) 1. Test Objective and Principle 1.1 Test Objective This test is based on Propionibacterium acnes and evaluates the acne-removing efficacy of the test sample by detecting its antibacterial effect on Propionibacterium acnes.

[0098] 1.2 Test Principle Acne is a chronic inflammatory skin disease that primarily affects the pilosebaceous unit. It is generally believed that acne is caused by four main factors: excessive sebum secretion, abnormal keratinization of the pilosebaceous duct, bacterial infection, and inflammatory response. Propionibacterium acnes is one of the main pathogens causing acne. This test establishes an antibacterial model against Propionibacterium acnes and evaluates the acne-reducing efficacy of the tested samples by detecting their antibacterial effect on Propionibacterium acnes.

[0099] 2. Test materials 2.1 Test System The bacterial strain used in this test was Propionibacterium acnes ATCC6919.

[0100] 2.2 Main Reagents BHIA, PBS.

[0101] 2.3 Main Equipment Biosafety cabinets and biochemical incubators.

[0102] 3. Testing Methods 1) The concentration tested in this experiment was 2% (diluted with PBS).

[0103] 2) Wash the fresh culture of the test bacteria with PBS and dilute it to a bacterial concentration of 5.0 x 10⁻⁶. 5 CFU / mL ~4.5x10 6 CFU / mL bacterial suspension.

[0104] 3) Take a sterile test tube, add 5.0 mL of sample, incubate in a 20℃±1℃ water bath for 5 min, then add 0.1 mL of the experimental bacterial suspension, mix quickly and start timing immediately. After 10 min, take a sample from the mixture of test bacteria and sample and dilute it. Select 2-3 dilutions for viable colony counting. Simultaneously, use PBS instead of the sample and repeat the above steps as a control sample. Calculate the inhibition rate using the following formula.

[0105] Antibacterial rate = (average colony count of control sample - average colony count of test sample) / average colony count of control sample × 100%.

[0106] 4. Test Results Table 2 Performance Test 3: Oil Control Efficacy (5α-Reductase Inhibition Rate) 1. Test Objective and Principle 1.1 Test Objective This test is based on 5α-reductase. By detecting the degree of inhibition of 5α-reductase activity by the sample, the oil-controlling efficacy of the sample is evaluated.

[0107] 1.2 Test Principle 5α-Reductase is a key enzyme in the metabolism of androgens in the skin, reducing testosterone to dihydrotestosterone (DHT). DHT is the most potent androgen receptor agonist. Excessive androgen secretion leads to hyperfunction of the sebaceous glands, stimulating them to produce more sebum. Inhibiting 5α-reductase activity effectively reduces DHT production and improves sebum secretion. A higher inhibition rate of 5α-reductase indicates a stronger oil-controlling effect, and vice versa. Based on this principle, this test uses the degree of inhibition of 5α-reductase activity by the experimental sample to determine the sample's oil-controlling efficacy.

[0108] 2. Test materials 2.1 Test System The enzyme used in this test was 5α-reductase.

[0109] 2.2 Main Reagents Phosphate buffer, NADPH, 5α-reductase, testosterone, dutazon.

[0110] 2.3 Main instruments and equipment: shaking plate, microplate reader, biochemical incubator.

[0111] 3. Testing Methods 1) The sample concentration in this experiment was 2% (diluted with phosphate buffer); positive control group: dutoxane 2.5 mmol / L; negative control: phosphate buffer.

[0112] 2) Determination of blank control: Add 4 μL of testosterone (20.747 mmol / L), 6 μL of 5α-reductase (176 mg / mL), 60 μL of phosphate buffer (pH = 7.4), and 30 μL of NADPH to a reaction tube. After mixing, measure the absorbance. Incubate at 37℃ for 60 min, and then measure the absorbance again. Subtract the background value of NADPH decrease to determine the blank decrease value (ΔA0). 3) Sample determination: Add 4 μL of testosterone T (20.747 mmol / L), 6 μL of 5α-reductase (176 mg / mL), 5 μL of the sample to be tested, 55 μL of phosphate buffer (pH = 7.4), and 30 μL of NADPH to the reaction tube. After mixing, measure the absorbance. Incubate at 37℃ for 60 min and measure the absorbance again. Subtract the background value of the NADPH blank and measure the decrease value of the sample (ΔA1).

[0113] 5α-reductase inhibition rate = (ΔA0 - ΔA1) / ΔA × 100%.

[0114] 4. Test Results Table 3 As can be seen from Tables 1-3, the plant-based oil-controlling and acne-removing compositions provided in Examples 1-3 of this invention have a significant inhibitory effect on the production of lipids in human immortalized sebaceous gland cells induced by sodium palmitate at a concentration of 0.2%, a significant antibacterial effect on Propionibacterium acnes at a concentration of 2%, indicating that they have an acne-removing effect; and a significant inhibitory effect on 5α-reductase at a concentration of 2%, indicating that they have an oil-controlling effect.

[0115] Compared with Example 3, Examples 4-6 show that the compound plant extract lacks any of the components of Saururus chinensis, Artemisia argyi, and Magnolia officinalis leaves, resulting in a significant decrease in the inhibitory effect of the plant-based oil-controlling and acne-removing composition on lipids in sebaceous gland cells, the antibacterial effect on Propionibacterium acnes, and the inhibitory effect on 5α-reductase. This indicates that Saururus chinensis, Artemisia argyi, and Magnolia officinalis leaves have a synergistic effect.

[0116] Compared with Example 3, Examples 7-11 show that the plant-based oil-controlling and acne-removing composition obtained by the preparation method of the compound plant extract provided by the present invention has significantly enhanced acne-removing and oil-controlling effects. Compared with Example 3, Examples 12-14 show that replacing the extract of *Gynostemma pentaphyllum* with Sophora flavescens extract, which has similar efficacy to that of the present invention, and replacing the extract of *Rhodiola rosea* with *Comfrey arvense* extract, resulted in a significant decrease in the inhibitory effect of the plant-based oil-controlling and acne-removing composition on lipids in sebaceous gland cells, the antibacterial effect on *Propionibacterium acnes*, and the inhibitory effect on 5α-reductase. The composition prepared using extracts of *Forsythia suspensa*, Sophora flavescens, *Salix matsudana*, and *Rhodiola rosea* with similar efficacy also had poor performance, indicating that the extracts of *Gynostemma pentaphyllum* and *Comfrey arvense*, together with the compound plant extracts, have a synergistic effect.

[0117] Performance Test 4: Clinical Verification of Oil Control Efficacy 1. Test Basis [1]T / ZHCA002-2018 Test Method for Oil Control Efficacy of Cosmetics. [2] Liu Weiyi, Zhou Lin, Zhao Hua. Cosmetic efficacy evaluation (XII) - consumer use test [J]. Daily Chemical Industry, 2021, 51(6): 485-490. [3] Laboratory protocol method. 2. Test Objective This test measured the changes in skin oil content and average oil spot size before and after using the test sample in 31 male or female subjects who met the criteria of oily facial skin and forehead oil content >120ug / cm2 (data before washing face). This test verified the claims of the submitted sample in oil control efficacy.

[0118] 3. Test Plan 3.1. Sample Information The plant-based oil-controlling and acne-removing composition of Example 3 was added to the essence water at an amount of 1 wt%, and the formula is shown in Table 4 below.

[0119] Table 4 3.2. Sample Usage Method Application area: Face Usage frequency and cycle: Use in the morning and evening for 28 consecutive days.

[0120] How to use: Apply morning and evening. Squeeze an appropriate amount of essence into the palm of your hand and gently massage onto your face until absorbed.

[0121] 3.3. Subject Information Subject gender: male or female Subject age: 19-45 years old Number of participants included: 32 Number of people who failed to complete the test: 1 Valid participants in data analysis: 31 Note: Since No. 02 withdrew, they are not included in the statistics.

[0122] 3.4. Inclusion and Exclusion Criteria for Subjects Participants in this trial were selected from the Noah Efficacy Laboratory's participant information database, choosing healthy participants who met the following inclusion and exclusion criteria: 3.4.1. Selection Criteria Healthy males or females aged 18-45; Oily facial skin, with an oil content on the forehead >120ug / cm2 (data before washing face); They can cooperate well with the test subjects and maintain a regular lifestyle during the research period; Able to read and understand all contents of the informed consent form, and willing to sign the informed consent form; During the testing period, participants agree not to use any cosmetics, drugs, or health products that may affect the results. Products with the same efficacy as the test samples must not be used during the testing period; During the testing period, no other cosmetics or personal care products may be used except for the test sample; Currently, I am not involved in, and will not be involved in, any other research involving the test area during the research period; No cosmetic procedures may be performed during the testing period.

[0123] 3.4.2. Exclusion Criteria Anyone meeting any of the following conditions must be excluded from this study. Those with skin conditions that may affect their judgment of test results; People with a high risk of allergies; Women who are pregnant, breastfeeding, or planning to become pregnant during the testing period; Those with severe heart, liver, or kidney dysfunction and severe immunodeficiency; Those with mental illness, severe endocrine disorders, or those taking oral contraceptives; Those who participated in drug clinical trials or other tests within the past 30 days, or those who have systematically used drugs that may affect test results within the past 2 weeks; Individuals who have used oral or topical beauty products within the past two weeks that may affect the test results; or individuals deemed unsuitable for this study by the researchers.

[0124] 3.4.3. Exit Criteria Patients may withdraw from a clinical study due to complications, adverse events, or other reasons during the study period, but the reason for withdrawal should be stated.

[0125] 3.5. Environmental conditions The test environment conditions were controlled throughout the entire testing process: temperature 21±1°C, humidity 50±10%RH.

[0126] 3.6. Sample Preparation Before testing, laboratory technicians blinded the samples and removed vendor-related information from the sample packaging to avoid affecting the test results. After testing, all test samples were collected.

[0127] 3.7. Data Analysis 3.7.1. Descriptive Statistics Statistical analysis software is used to perform statistical analysis on the data, calculating the mean, standard deviation, and rate of change of each parameter at various time points. Calculation formula: Rate of change = [(Value after use - Value before use) / Value before use] x 100% 3.7.2. Normality Test If the normality test of the above data is performed and the asymptotic significance (two-tailed) value is >0.05, then the data series follows a normal distribution.

[0128] 3.7.3. Difference Analysis Using before-and-after sample comparisons: When both sets of data are normally distributed, use a paired T-test to analyze the differences between the two sets of data; when the two sets of data are not normally distributed, use a rank-sum test to analyze the differences between the two sets of data.

[0129] 3.7.4. Evaluation Criteria "ns" indicates no significant difference, P ≥ 0.05; "Indicates a significant difference, 0.010 ≤ P < 0.05;" "This indicates a significant difference, 0.001 ≤ P < 0.010;" "Indicates a significant difference, P<0.001.

[0130] 3.8. Criteria for Result Judgment If the skin's oil content or average oil spot size significantly decreased before and after using the sample (P<0.05), then the test sample is considered to have oil-controlling effects.

[0131] 3.9. Test Results Table 5 Compared with before sample use, after 14 days of sample use, the skin oil content of the 31 subjects showed a significant difference, with a change rate of -19.15% compared with the baseline value; after 28 days of sample use, the skin oil content showed a significant difference, with a change rate of -32.22% compared with the baseline value. Compared with before sample use, after 14 days of sample use, the average oil spot size of the 31 subjects was significantly different, with a change rate of -30.63% compared with the baseline value; after 28 days of sample use, the average oil spot size was significantly different, with a change rate of -53.71% compared with the baseline value.

[0132] 4. Test Conclusion Under the test conditions, the test sample showed oil-controlling effects after 14 and 28 days of use.

[0133] VISIA images of some subjects at different time points - standard as follows Figure 1 As shown; VISIA images of some subjects at different time points - porphyrins Figure 2 As shown.

[0134] The above description represents the preferred embodiments of the present invention. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principles of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A plant-based oil-controlling and acne-removing composition, characterized in that, By weight percentage, it includes 3-5% compound plant extract, 2-3% goldenrod extract, 1-2% comfrey extract, and solvent to make up to 100%; The plant raw materials for the compound plant extract consist of Artemisia argyi, Saururus chinensis and Magnolia officinalis leaves in a mass ratio of 4-6:2-3:2-3. The preparation method of the compound plant extract is as follows: S1. Crush and sieve the dried plant material to obtain raw material powder; S2. The raw material powder is extracted sequentially using supramolecular solvent extraction, enzymatic hydrolysis with compound enzyme preparation, and extraction with octanol-thymol solution containing phosphatidylcholine. The extracts are combined, concentrated and dried under reduced pressure to obtain the compound plant extract. The supramolecular solvent is an organic acid-hydroxytyrosol aqueous solvent.

2. The plant-based oil-controlling and acne-removing composition according to claim 1, characterized in that, The supramolecular solvent is prepared by mixing an organic acid, hydroxytyrosol, and water, heating to 50-60°C, and reacting under nitrogen protection for 2-4 hours.

3. The plant-based oil-controlling and acne-removing composition according to claim 2, characterized in that, The molar ratio of the organic acid to hydroxytyrosol is 1-3:1-3; the water content of the supramolecular solvent is 10-20 wt%.

4. The plant-based oil-controlling and acne-removing composition according to claim 3, characterized in that, The organic acid is selected from at least one of citric acid, lactic acid, malic acid, and succinic acid.

5. The plant-based oil-controlling and acne-removing composition according to any one of claims 1-4, characterized in that, The compound enzyme preparation consists of cellulase and pectinase in a mass ratio of 1:0.3-0.

5.

6. The plant-based oil-controlling and acne-removing composition according to any one of claims 1-4, characterized in that, The method for preparing the octanol-thymol solution containing phosphatidylcholine is as follows: mix octanol and thymol, stir at 70-80℃ for 2-4 hours, add phosphatidylcholine and mix evenly to obtain the solution.

7. The plant-based oil-controlling and acne-removing composition according to claim 6, characterized in that, The molar ratio of octanol to thymol is 1:2-4; the octanol-thymol solution containing phosphatidylcholine contains 2-4 wt% phosphatidylcholine.

8. The plant-based oil-controlling and acne-removing composition according to any one of claims 1-4, characterized in that, Step S2 is as follows: Mix the raw material powder and supramolecular solvent, extract by ultrasound, add the compound enzyme preparation, hydrolyze by enzymatic hydrolysis, inactivate the enzyme by ultrasound, filter, and obtain the first extract and filter residue; mix the filter residue with an octanol-thymol solution containing phosphatidylcholine, stir and extract, filter, and obtain the second extract; combine the first extract and the second extract, concentrate and dry under reduced pressure to obtain the compound plant extract.

9. A method for preparing the plant-based oil-controlling and acne-removing composition according to any one of claims 1-8, characterized in that, Includes the following steps: The compound plant extract, yellow clover extract, comfrey extract and solvent are mixed evenly to obtain the final product.

10. The use of the plant-based oil-controlling and acne-removing composition according to any one of claims 1-8 in the preparation of oil-controlling and acne-removing products.

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