Composition for controlling oil, shrinking pores and tightening skin as well as preparation method and application of composition

Abstract

The invention relates to a composition for controlling oil, shrinking pores and tightening skin as well as a preparation method and application thereof, and belongs to the technical field of cosmetics. The composition with the effects of controlling oil, shrinking pores and tightening skin, provided by the invention, is prepared from EGCG (epigallocatechin gallate) nanocrystals, a phellinus igniarius extract, a lactobacillus / pomegranate fruit fermentation product and cardamom seed ketone, the EGCG nanocrystal is prepared from an EGCG solution in a micro-channel reactor through an anti-solvent crystallization method by taking supercritical CO2 as an anti-solvent. The composition synchronously inhibits the activity of 5alpha-reductase, regulates sebaceous gland secretion, enhances keratinocyte metabolism and promotes barrier repair through a multi-target synergistic mechanism, and improves the stability and transdermal absorption efficiency of active ingredients by means of a specific fermentation process. Therefore, the long-acting and mild oil control, tightening and pore repairing effects with skin health promotion significance are realized.

Description

Technical Field

[0001] This invention relates to the field of cosmetic technology, and in particular to a composition for controlling oil, shrinking pores and firming skin, its preparation method and application. Background Technology

[0002] Currently, to achieve oil control, astringent, and firming effects, similar products or methods mainly rely on chemical exfoliants such as salicylic acid and fruit acids, or simple combinations of single plant extracts (such as witch hazel and tea tree oil). These solutions achieve short-term apparent improvement by dissolving keratin, providing temporary physical astringency, or offering mild antibacterial effects. However, they generally suffer from the following technical drawbacks: a single and superficial mechanism of action, failing to regulate sebaceous gland activity at its source, leading to rebound after discontinuation; significant irritation with long-term use, as chemical exfoliation can damage the skin barrier, causing dryness, sensitivity, and even compensatory oil production; insufficient ingredient stability and bioavailability, making it difficult for some active ingredients to effectively penetrate to the target area; and a lack of comprehensive regulation of the skin's microecology and inflammatory pathways, failing to address the common microbial imbalances and chronic mild inflammation associated with oily skin, thus only treating the symptoms, not the root cause. Furthermore, highly effective active ingredients such as EGCG suffer from poor stability, low transdermal absorption, and limited compatibility, making it difficult to fully realize their multi-pathway regulatory potential. Therefore, there is an urgent need to improve the problems of poor stability, low transdermal absorption rate and limited compatibility of highly effective active ingredients, and to provide a product that achieves oil control, astringent and firming effects through multiple mechanisms of deep action. Summary of the Invention

[0003] The purpose of this invention is to overcome the shortcomings of the prior art and provide a composition for controlling oil, shrinking pores and tightening skin, as well as its preparation method and application.

[0004] To achieve the above objectives, the technical solution adopted by the present invention is as follows: In a first aspect, the present invention provides a composition for controlling oil, shrinking pores and tightening skin, comprising the following components in parts by weight: 0.01-0.5 parts of EGCG nanocrystals, 0.01-4 parts of medicinal blastocystis extract, 0.01-2.5 parts of Lactobacillus / pomegranate fermentation product, and 0.001-0.1 parts of cardamom seed ketone; wherein the EGCG nanocrystals are prepared by antisolvent crystallization of EGCG solution in a microchannel reactor using supercritical CO2.

[0005] First, this invention prepares EGCG nanocrystals via an antisolvent crystallization method, significantly improving the bioavailability and efficacy stability of EGCG. The EGCG nanocrystals are spherical or near-spherical particles with good dispersibility and a relatively narrow particle size distribution, with an average particle size of 150-225 nm. By increasing specific surface area and surface energy, EGCG nanocrystals greatly enhance their solubility and transdermal absorption rate. Compared to ordinary EGCG, nanocrystal technology achieves three major advantages: high transdermal absorption, strong targeted delivery, and long-lasting stability, making the effects of oil control, pore shrinking, and skin tightening more significant and lasting.

[0006] Furthermore, the composition achieves comprehensive effects of oil control, pore tightening, and skin firming through a multi-pathway synergistic mechanism. Specifically, EGCG nanocrystals, as the core ingredient, reduce excessive sebum secretion from the source by inhibiting 5α-reductase activity. Simultaneously, their strong antioxidant capacity scavenge free radicals, reducing oxidative stress on the sebaceous glands. The medicinal extract of *Polyporus thuringiensis* is rich in triterpenoids, which can tighten pores, regulate water-oil balance, and have anti-inflammatory effects, improving inflammatory responses caused by excessive sebum secretion. Lactobacillus / pomegranate fermentation products regulate the skin's microecological balance, inhibit the proliferation of harmful bacteria, and promote the colonization of beneficial bacteria. At the same time, the organic acids and active small molecule peptides produced by the fermentation process gently promote keratin renewal and enhance the skin barrier function. Cardamom seed ketone, by inhibiting sebaceous gland cell differentiation and reducing sebum synthesis, synergistically enhances the oil control effect. Through scientific formulation, these ingredients form a multi-dimensional synergistic network of "inhibiting sebum secretion - tightening pores - regulating microecology - repairing the skin barrier." Compared with single-ingredient or traditional acid products, the composition provided by this invention has significant advantages such as comprehensive mechanism of action, strong oil control and long-lasting effect, low irritation and repair function, achieving a gentle and long-lasting oil control and firming effect.

[0007] Further, the composition comprises the following components in parts by weight: 0.05-0.3 parts of EGCG nanocrystals, 0.1-3 parts of medicinal blastocystis extract, 0.1-1 parts of Lactobacillus / pomegranate fruit fermentation product, and 0.008-0.05 parts of cardamom seed ketone.

[0008] More preferably, the composition comprises the following components in parts by weight: 0.08-0.1 parts of EGCG nanocrystals, 0.5-1 parts of medicinal blastocystis extract, 0.4-0.6 parts of Lactobacillus / pomegranate fermentation product, and 0.01-0.03 parts of cardamom seed ketone.

[0009] Furthermore, the preparation method of the EGCG nanocrystals includes the following steps: S1. Dissolve EGCG in ethyl acetate, stir and sonicate under light-protected conditions, filter, collect the filtrate, and obtain an EGCG solution; S2. Using supercritical CO2 as the antisolvent, supercritical CO2 and EGCG solution were passed into a microchannel reactor for antisolvent crystallization to prepare EGCG nanocrystals.

[0010] Preferably, the concentration of the EGCG solution in step S1 is 10-15 mg / mL.

[0011] Preferably, the stirring in step S1 is magnetic stirring at a speed of 300-400 r / min for 15-25 min; the ultrasonic treatment is ultrasonication at an ultrasonic power of 90-100 W for 20-30 min.

[0012] Preferably, the filtration in step S1 is performed using an organic phase filter with a pore size of 0.22-0.45 μm.

[0013] Preferably, the antisolvent crystallization conditions in step S2 are as follows: at 35-37°C and 10-12 MPa pressure, the flow rate of supercritical CO2 is controlled at 33-42 mL / min and the flow rate of EGCG solution is controlled at 1-2 mL / min.

[0014] Further, the preparation method of the Lactobacillus / pomegranate fruit fermentation product is as follows: after washing the mature pomegranate berries, add 5-10 times the weight of water, crush and pulp them to obtain pomegranate pulp, and sterilize it; inoculate the sterilized pomegranate pulp with Lactobacillus rhamnosus culture, and anaerobic ferment at 36-38℃ for 60-84h to obtain the fermentation product; centrifuge and filter the fermentation product to remove bacteria, obtain the fermentation broth, and freeze-dry it under vacuum to obtain the Lactobacillus / pomegranate fruit fermentation product.

[0015] Preferably, the inoculation amount of the *Lactobacillus rhamnosus* culture is 3% v / v-6% v / v of pomegranate pulp, and the viable count of the *Lactobacillus rhamnosus* culture is 2 × 10⁻⁶. 8 -3×10 8 CFU / mL.

[0016] Secondly, the present invention provides the use of the composition described in the first aspect in the preparation of cosmetics having the effects of oil control, pore shrinking and skin tightening.

[0017] Furthermore, the cosmetics include one of the following: toner, lotion, cream, mask, and spray.

[0018] Thirdly, the present invention provides an oil-controlling, pore-minimizing, and skin-tightening essence lotion, comprising the following components in weight percentages: 0.05%-0.5% thickener, 1%-10% moisturizer, 0.01%-0.3% pH adjuster, 1%-6% emulsifier, 5%-15% oil, 0.5%-3% preservative, 0.01%-0.3% chelating agent, 1%-5% of the composition described in the first aspect, and the balance being deionized water.

[0019] Furthermore, the said essence lotion includes at least one of the following (a)-(e): (a) The thickener comprises at least one of xanthan gum, carbomer, hydroxyethyl acrylate / sodium acryloyl dimethyl taurate copolymer, ammonium acryloyl dimethyl taurate / VP copolymer, hydroxyethyl cellulose, and cetyl alcohol; (b) The moisturizer includes at least one of allantoin, sodium polyacrylate, panthenol, beta-glucan, trehalose, caprylyl glycol, dipropylene glycol, sodium hyaluronate, 1,2-butanediol, glycerin, and tremella polysaccharide; (c) The pH adjuster includes at least one of arginine, citric acid, and NaOH; (d) The emulsifier includes at least one of PEG-100 glyceryl stearate, glyceryl stearate citrate, cetearyl glucoside, cetearyl alcohol, PEG-20 methyl glucose sesquistearate, methyl glucose sesquistearate, cetyl phosphate potassium, and sodium stearoyl glutamate; (e) The oils include at least one of caprylic / capric triglyceride, polydimethylsiloxane, jojoba oil, grape seed oil, meadowfoam seed oil, squalane, macadamia nut oil, and camellia oil.

[0020] (f) The preservative includes at least one of 1,2-hexanediol, 1,2-pentanediol, ethylhexylglycerin, p-hydroxyacetophenone, capryloyl hydroxamic acid, and sodium benzoate.

[0021] (g) The chelating agent includes EDTA-2Na.

[0022] Compared with the prior art, the beneficial effects of the present invention are as follows: This invention is based on the scientific relationship between skin lipid metabolism, microecological homeostasis, and epidermal barrier integrity. Combining modern fermentation engineering technology with research findings on natural active ingredients, it constructs a multi-effect compound system consisting of EGCG nanocrystals, medicinal *Polyporus thuringiensis* extract, *Lactobacillus* / pomegranate fermentation products, and cardamom seed ketone. This composition, through a multi-target synergistic mechanism, simultaneously inhibits 5α-reductase activity, regulates sebaceous gland secretion, enhances keratinocyte metabolism, and promotes barrier repair. Furthermore, a specific fermentation process enhances the stability of the active ingredients and transdermal absorption efficiency, thereby achieving long-lasting, gentle oil control, firming, and pore-repairing effects that promote skin health. Detailed Implementation

[0023] To better illustrate the purpose, technical solution, and advantages of the present invention, the present invention will be further described below in conjunction with specific embodiments.

[0024] In the following embodiments, the source or preparation method of the raw materials is as follows: EGCG: Product No.: B20106, Manufacturer: Shanghai Yuanye; Cardamom Seed Ketone: Product Code: SymDecanox TM DPG, Manufacturer: Symrise; Medicinal Fomitopsis pilosula extract: Product No.: Bio-Fomes TM Manufacturer: Dongfang Miaosen; The preparation method of Lactobacillus / pomegranate fruit fermentation product is as follows: After washing ripe pomegranate berries, add 8 times their weight of water and crush and pulp them to obtain ripe pomegranate pulp. Sterilize the pulp in a water bath at 100℃ for 30 minutes. Inoculate the sterilized pomegranate pulp with 4.5% v / v and a viable count of 2.5 × 10⁻⁶ cells / ml. 8 A CFU / mL Lactobacillus rhamnosus culture (strain number CICC 6013, purchased from the China Industrial Microbial Culture Collection Center) was anaerobically fermented at 37°C for 72 h to obtain the fermentation product. The fermentation product was centrifuged at 4°C and 8000 r / m for 15 min, and the supernatant was filtered through a 0.22 μm filter to remove bacteria, yielding the fermentation broth. The fermentation broth was pre-frozen at -80°C for 4 h, and then transferred to a freeze dryer and freeze-dried according to the following program: pre-freezing at -40°C for 2 h, sublimation at -40°C to -20°C for 24 h, and desorption at -20°C to 25°C for 10 h, under a vacuum of 10 Pa to obtain the Lactobacillus / pomegranate fruit fermentation product.

[0025] Unless otherwise specified, all other materials and reagents used in the examples are commercially available.

[0026] Example 1 This embodiment provides an EGCG nanocrystal 1, the preparation method of which includes the following steps: S1. Dissolve EGCG in ethyl acetate to prepare a solution of 12.5 mg / mL. Stir magnetically at 350 r / min for 20 min in the dark, and then sonicate at 95 W for 25 min to ensure complete dissolution. Filter the solution through an organic phase filter (13 mm in diameter, 0.22 μm in pore size) and collect the filtrate to obtain the EGCG solution. S2. Using a microchannel reactor, under conditions of 36℃ and 11MPa pressure, supercritical CO2 was used as the antisolvent. The supercritical CO2 flow rate was controlled at 38mL / min and the EGCG solution feed rate was controlled at 1.5mL / min for antisolvent crystallization. The collected EGCG nanocrystal suspension was pre-frozen at -80℃ for 6 h and then transferred to a freeze dryer and freeze-dried according to the program: the temperature was increased stepwise from -40℃ to 25℃ at a heating rate of 2.5℃ / h and the vacuum degree was 10 Pa to obtain EGCG nanocrystals 1.

[0027] Example 2 This embodiment provides an EGCG nanocrystal 2, the preparation method of which includes the following steps: S1. Dissolve EGCG in ethyl acetate to prepare a 10 mg / mL solution. Stir magnetically at 300 r / min for 25 min in the dark, and then sonicate at 90 W for 30 min to ensure complete dissolution. Filter the solution through an organic phase filter (13 mm in diameter, 0.22 μm in pore size) and collect the filtrate to obtain the EGCG solution. S2. Using a microchannel reactor, under conditions of 35℃ and 12MPa pressure, supercritical CO2 was used as the antisolvent. The supercritical CO2 flow rate was controlled at 33mL / min and the EGCG solution inlet rate was 1mL / min for antisolvent crystallization. The collected EGCG nanocrystal suspension was pre-frozen at -80℃ for 6 h and then transferred to a freeze dryer and freeze-dried according to the program: the temperature was increased stepwise from -40℃ to 25℃ at a heating rate of 2.5℃ / h and the vacuum degree was 10 Pa to obtain EGCG nanocrystals 2.

[0028] Example 3 This embodiment provides an EGCG nanocrystal 2, the preparation method of which includes the following steps: S1. Dissolve EGCG in ethyl acetate to prepare a 15 mg / mL solution. Stir magnetically at 400 r / min for 15 min in the dark, and then sonicate at 100 W for 20 min to ensure complete dissolution. Filter the solution through an organic phase filter (13 mm in diameter, 0.22 μm in pore size) and collect the filtrate to obtain the EGCG solution. S2. Using a microchannel reactor, under conditions of 37℃ and 10MPa pressure, supercritical CO2 was used as the antisolvent. The supercritical CO2 flow rate was controlled at 42mL / min and the EGCG solution inlet rate was controlled at 2mL / min for antisolvent crystallization. The collected EGCG nanocrystal suspension was pre-frozen at -80℃ for 6 h and then transferred to a freeze dryer and freeze-dried according to the program: the temperature was increased stepwise from -40℃ to 25℃ at a heating rate of 2.5℃ / h and the vacuum degree was 10 Pa to obtain EGCG nanocrystals 3.

[0029] Comparative Example 1 This comparative example provides an EGCG nanocrystal 1', the preparation method of which differs from that of Example 1 in that: in step S2, the supercritical CO2 flow rate is 45 mL / min and the EGCG solution inlet rate is 0.5 mL / min; The remaining steps and parameters are the same as in Example 1, and EGCG nanocrystals 1' are prepared.

[0030] Comparative Example 2 This comparative example provides an EGCG nanocrystal 2', the preparation method of which differs from that of Example 1 in that: in step S2, the supercritical CO2 flow rate is 30 mL / min and the EGCG solution inlet rate is 3 mL / min; The remaining steps and parameters are the same as in Example 1, and EGCG nanocrystals 2' are prepared.

[0031] Comparative Example 3 This comparative example provides an EGCG nanocrystal 3', the preparation method of which differs from that of Example 1 in that the pressure in step S2 is 15 MPa; The remaining steps and parameters are the same as in Example 1, and EGCG nanocrystals 3' are prepared.

[0032] Comparative Example 4 This comparative example provides an EGCG nanocrystal 4', the preparation method of which differs from that of Example 1 in that the pressure in step S2 is 8 MPa; The remaining steps and parameters are the same as in Example 1, and EGCG nanocrystals 4' are prepared.

[0033] Comparative Example 5 This comparative example provides an EGCG nanocrystal 5', the preparation method of which differs from that of Example 1 in that the temperature in step S2 is 40℃; The remaining steps and parameters are the same as in Example 1, and EGCG nanocrystals 5' are prepared.

[0034] Comparative Example 6 This comparative example provides an EGCG nanocrystal 6', the preparation method of which differs from that of Example 1 in that the temperature in step S2 is 30°C; The remaining steps and parameters are the same as in Example 1, and EGCG nanocrystals 6' are prepared.

[0035] Test Example 1: Particle size test and skin penetration performance test of EGCG nanocrystals Test samples: EGCG nanocrystals prepared in Examples 1-3 and Comparative Examples 1-6.

[0036] (1) Particle size test: The particle size of EGCG nanocrystals was determined using dynamic light scattering (DLS). Specifically, 200 μL of an aqueous solution of EGCG nanocrystals (concentration 1.0 μg / mL) was diluted with 1 mL of PBS buffer and thoroughly mixed. A nanoparticle size analyzer (WINNER-801) was used, with the detection angle set to 90°. A photon correlator received the light signal. Detection conditions were as follows: temperature: 25℃ (thermostatic control); equilibration time: 120 seconds (to ensure sample temperature stability); measurements were repeated three times for each sample, with 300 data points measured each time to improve statistical accuracy. The average particle size and polydispersity index (PDI) were automatically calculated using software. The average particle size results are shown in Table 1.

[0037] (2) Skin permeability test Experimental method: Ex vivo pig skin was used for testing, and the exposed skin area in the diffusion cell was 1.5 cm². 2 The receiving chamber had a volume of 5.0 mL. A 50:50 volume mixture of phosphate buffer (pH 5.8) and anhydrous ethanol was used as the receiving solution. 500 μL of each of the 10 wt% EGCG nanocrystal solution samples (solvent: 1,3-propanediol) were evenly applied to the skin (10 wt% EGCG solution served as the control group, solvent: 1,3-propanediol), and covered with a thin film to prevent evaporation. The transdermal absorption of EGCG nanocrystals at different time points was measured by HPLC, and the cumulative permeation after 48 h (in μg / cm³) was recorded and calculated. 2 The formula for calculating the cumulative infiltration at the nth time point is as follows: ; In the formula: Qn: Cumulative permeability at the nth time point (unit: μg / cm³) 2 ); Vr: Receiving chamber volume (i.e., total volume of receiving liquid, Vr = 5.0 mL); Cn: The concentration of EGCG in the receiving solution measured at the nth time point (unit: μg / mL). Vs: Sampling volume per sample (unit: mL, here it is 0.5 mL); Ci: The concentration of EGCG measured in the receiving solution at the i-th time point (unit: μg / mL). Correction is performed on the total amount of EGCG extracted in the first n-1 samplings (unit: μg). A: The area of ​​skin exposed in the diffusion pool (A=1.5cm²) 2 ).

[0038] The greater the cumulative amount of penetration, the better the skin's permeability.

[0039] The EGCG nanocrystals prepared in Examples 1-3 were spherical or near-spherical particles. The average particle size was measured as shown in Table 1. The EGCG nanocrystals prepared in Examples 1-3 had a relatively narrow particle size distribution and good dispersibility, with an average particle size range of 150-225 nm. From the data of Comparative Examples 1-6, it can be seen that the flow rate ratio of supercritical CO2 and EGCG solution, as well as the temperature and pressure settings in the microchannel reactor during the preparation of EGCG nanocrystals, have a significant impact on the average particle size of the prepared EGCG nanocrystals.

[0040] As shown in Table 1, the skin permeability test results indicate that the cumulative permeability of the EGCG nanocrystals prepared in Examples 1-3 was significantly increased compared to untreated EGCG, reaching 168-211 μg / cm³. 2 This indicates that the transdermal absorption performance of nanocrystalline EGCG is greatly improved compared to untreated EGCG. Comparing the cumulative permeation data of Comparative Examples 1-6 with that of Example 1, the cumulative permeation of EGCG nanocrystals in Example 1 is much higher than that in Comparative Examples 1-6, indicating that the step parameters in the preparation process of EGCG nanocrystals also have a significant impact on their final transdermal absorption performance.

[0041] Table 1. Particle size test and skin penetration performance test of EGCG nanocrystals Group Average particle size (nm) <![CDATA[Penetration cumulative amount (μg / cm 2 )]]> Example 1 201 211 Example 2 223 168 Example 3 186 185 Comparative Example 1 150 108 Comparative Example 2 235 123 Comparative Example 3 265 95 Comparative Example 4 511 31 Comparative Example 5 240 112 Comparative Example 6 296 103 control group - 26 Application examples and comparative application examples This invention provides a composition for controlling oil, shrinking pores, and tightening skin. It uses EGCG nanocrystals 1 prepared in Example 1, combined with medicinal Ferula melitica extract, Lactobacillus / pomegranate fermentation product, and cardamom seed ketone. Specifically, compositions 1-7 of the application examples and compositions 1'-5' of the comparative application examples are prepared according to the formulas shown in Table 2.

[0042] Table 2. Component composition and dosage of the composition

[0043] Note: "-" indicates no addition, and the total weight of compositions 1-7 and compositions 1'-5' is the same.

[0044] Furthermore, the present invention also provides an essence lotion that controls oil, shrinks pores, and tightens skin. Specifically, according to the formula shown in Table 3, the obtained compositions 1-7 and compositions 1'-5' are respectively prepared into essence lotions to obtain essence lotions 1-9, essence lotions 1'-5', and blank essence lotions without the addition of compositions. Among them: lotions 1-7 are prepared by adding compositions 1-7 in sequence, such as lotion 1 is prepared by adding composition 1, and so on; lotions 8 and 9 are prepared by adding composition 4; lotion 1' is prepared by adding composition 1', and so on.

[0045] The preparation method of the essence emulsion includes the following steps: (1) Mix the thickener and humectant evenly, add deionized water, heat at 75°C for 10 minutes, then homogenize in a homogenizer at 4000 r / min for 5 minutes, keep warm at 75°C for later use, and obtain the pre-prepared component A.

[0046] (2) Take the oil and emulsifier in a beaker, heat to 75°C, stir until uniform, keep warm at 75°C for later use, and obtain the pre-made component B.

[0047] (3) In a homogenizer at 75℃ and 4000r / min, the pre-made component B is added to the pre-made component A and homogenized for 5 minutes to obtain the emulsified base material.

[0048] (4) Stir the emulsified base material and cool it down to 50°C. Add each component of the composition and continue stirring until the material is uniform. Then add the preservative and chelating agent, stir evenly, adjust the pH with a pH adjuster, stop stirring, discharge the material, and obtain the essence emulsion.

[0049] Table 3. Ingredients and dosage of the serum lotion (weight percentage, %)

[0050] Test Example 2: In vitro oil-controlling, antioxidant, and skin-firming effects of the composition. Using compositions 1-7 and 1'-5' as test samples, in vitro tests were conducted to assess the oil-controlling, antioxidant, and skin-firming effects of the compositions. The test results are as follows: 1. In vitro 5-α reductase inhibition effect test Experimental methods: (1) Reagent preparation Sample solutions: Compositions 1-7 and Compositions 1'-5' were each prepared with pure water to form sample solutions with a mass fraction of 10% for later use; Enzyme solution: 5-α reductase was diluted with PBS buffer at pH 6.8 to prepare a 50U enzyme solution; NADPH solution: Prepare a 2mM NADPH solution using PBS at pH 6.8; Testosterone solution: A 0.83 nM testosterone solution prepared by using a 75% ethanol solution.

[0051] (2) Mixed reaction (a) Sample tubes: Add 1 mL of sample solution, 1 mL of enzyme solution, 1 mL of NADPH solution and 1 mL of testosterone solution to a test tube, shake gently to obtain a mixed liquid, and use a pipette to transfer 200 μL of the mixed liquid into a 96-well microplate. Perform three replicates for each sample. Place the plate in a microplate reader and measure the absorbance at 340 nm. This is the first measurement value, A sample 0. After incubating the sample at 37°C for 20 minutes, place it in a microplate reader and measure the absorbance at 340 nm. This is the second measurement value, A sample 1. (b) Enzyme tubes: Add 1 mL of pure water, 1 mL of enzyme solution, 1 mL of NADPH solution and 1 mL of testosterone solution to a test tube, shake gently to obtain a mixed liquid, and use a pipette to take 200 μL of the mixed liquid into a 96-well microplate. Perform three replicates for each sample, and place them in a microplate reader for detection. Measure the absorbance at 340 nm, which is the first measurement value, Aenzyme 0. After incubating the sample at 37°C for 20 minutes, place it in a microplate reader for detection. Measure the absorbance at 340 nm, which is the second measurement value, Aenzyme 1. At the same time, a blank control group is set up: replace the enzyme solution in the sample tube with an equal volume of pure water, which is used for zeroing during the measurement to subtract absorbance changes caused by non-enzymatic reactions in the reaction system (such as spontaneous substrate degradation, solvent background absorption, etc.) to ensure the accuracy of enzyme activity measurement.

[0052] (3) Data calculation and result analysis Formula for calculating 5-α reductase inhibition rate: 5-α reductase inhibition rate (%) = [1 - (A sample 0 - A sample 1) / (A enzyme 0 - A enzyme 1)] × 100%; The higher the 5-α reductase inhibition rate, the stronger the ability of the composition to block the conversion of testosterone to dihydrotestosterone, and the less oil secreted by the sebaceous glands.

[0053] 2. In vitro antioxidant assay (DPPH assay) (1) Reagent preparation: Preparation of DPPH solution: Weigh 4 mg of DPPH powder, dissolve it in 60% v / v ethanol aqueous solution, and make up to 100 mL to prepare a DPPH solution with a concentration of 40 mg / L. Store in the dark. Sample solution preparation: Using compositions 1-7 and 1'-5' as samples, dissolve the corresponding samples in 60% v / v ethanol aqueous solution and make up to volume to prepare a sample solution with a concentration of 5 mg / mL; (2) Experimental Groups: Blank control group (A0): 2 mL sample solution + 2 mL 60% v / v ethanol aqueous solution; Sample group (A1): 2 mL sample solution + 2 mL DPPH solution; Sample matrix group (A2): 2 mL DPPH solution + 2 mL 60% v / v ethanol aqueous solution; Reaction: After mixing the groups evenly, let them stand in the dark for 30 min, centrifuge at 5000 r / min for 10 min, take the supernatant, and measure the absorbance at a wavelength of 517 nm.

[0054] (3) Parallel experiments and replication: Three parallel tubes were set up for each sample, and the average value of the results was taken.

[0055] (4) Data calculation and result analysis DPPH radical scavenging rate formula: DPPH radical scavenging rate (%) = [1 − (A1 − A0) / A2] × 100%; A higher DPPH free radical scavenging rate indicates a better antioxidant effect.

[0056] 3. In vitro type III collagen content test (1) Cell culture and inoculation Human fibroblasts were seeded in T75 culture flasks and cultured in DMEM medium containing 10% v / v FBS at 37°C and 5% v / v CO2. When the cells reached the logarithmic growth phase, they were digested with 0.25% trypsin-EDTA solution, and the cell density was adjusted to 2 × 10⁶ cells / year. 5 Cells / mL were seeded into 96-well plates, with an appropriate amount of cell suspension added to each well (100 μL per well).

[0057] (2) Preparation of test samples Take composition 1-7 and composition 1'-5' respectively, mix and dilute with 1 mL of DMEM medium to obtain test sample 1-7 and test sample 1'-5' with a composition concentration of 100 μg / mL.

[0058] (3) Group setup and drug administration Control group: 100 μL cell suspension + 100 μL blank DMEM medium; Sample set: 100 μL cell suspension + 50 μL blank DMEM culture medium + 50 μL sample to be tested; Specific procedures: In a 96-well plate, administer the drug according to the group settings described above, so that the final volume reaches the culture volume of 200 μL.

[0059] (4) Cell culture and treatment The treated cells were incubated at 37°C and 5% v / v CO2 for 24 hours to observe the effect of the samples on type III collagen production.

[0060] (5) Sample collection and collagen detection Cell lysis: After culture, discard the culture medium and gently wash the cells twice with PBS. Add an appropriate amount of cell lysis buffer and lyse the cells on ice for 30 minutes, gently shaking occasionally to ensure complete lysis. After lysis, transfer the lysis buffer to a centrifuge tube, centrifuge at 12,000 rpm for 10 minutes at 4°C, and collect the supernatant.

[0061] Collagen detection: Follow the instructions for the Type III collagen detection kit, mix the supernatant with the reagents in the kit, and allow the reaction to proceed. Measure the absorbance of the reaction product using a microplate reader, and calculate the type III collagen content based on the standard curve.

[0062] The rate of change in type III collagen for each composition was calculated using the following formula: Type III collagen increase rate (%) = (sample group - control group) / control group × 100%; The higher the increase rate of type III collagen, the better the skin-firming effect.

[0063] The results are shown in Table 4. Compositions 1-7 all showed high 5-α reductase inhibition rate, DPPH free radical scavenging rate, and type III collagen increase rate, indicating that the in vitro experiments verified that compositions 1-7 provided by the present invention have better oil control, antioxidant, and skin firming effects. Comparing the data of compositions 1'-5' and composition 4: The difference between compositions 1'-4' and composition 4 lies in the absence of EGCG nanocrystals, medicinal blastocystis extract, Lactobacillus / pomegranate fermentation product, or cardamom seed ketone, respectively. The 5-α reductase inhibition rate, DPPH free radical scavenging rate, and type III collagen increase rate of compositions 1'-4' are significantly lower than those of composition 2, especially composition 1'. This indicates that the combination of EGCG nanocrystals, medicinal blastocystis extract, Lactobacillus / pomegranate fermentation product, and cardamom seed ketone in the composition can synergistically improve the oil control, antioxidant, and skin firming effects of the composition, with EGCG nanocrystals having a more significant impact on the composition's effects. The difference between composition 5' and composition 4 lies in the different amounts of each component in the composition, indicating that the ratio of each component in the composition also has an important influence on the oil control, antioxidant, and skin firming effects of the composition.

[0064] Table 4 Results of in vitro oil control, antioxidant, and skin firming effects tests Group 5-α reductase inhibition rate / % DPPH free radical scavenging rate / % Type III collagen increase rate / % Composition 1 87.9 85.3 148.3 Composition 2 91.0 87.4 151.7 Composition 3 93.8 92.8 155.1 Composition 4 96.3 95.1 156.9 Composition 5 95.2 90.5 153.5 Composition 6 88.5 86.2 149.7 Composition 7 85.4 81.7 145.2 Composition 1' 63.2 59.3 120.8 Composition 2' 64.8 62.8 127.6 Composition 3' 69.7 61.4 123.0 Composition 4' 67.1 65.6 124.9 Composition 5' 72.6 67.9 130.4 Test Example 3: Human Body Patch Test Thirty participants were selected for the test according to the inclusion criteria, and the sample size was no more than 50 mm². 2 A qualified spot test apparatus with a depth of approximately 1 mm is used. The sample is placed in the small chamber of the spot test apparatus, with a volume of approximately 0.020 mL to 0.025 mL. The spot test apparatus containing the sample (Essence Emulsion 1-9 and Essence Emulsion 1'-5', blank essence emulsion, and distilled water as the blank control) is applied to the flexor side of the subject's forearm using hypoallergenic adhesive tape. Gently press with the palm to ensure even application to the skin, and leave for 24 hours. Skin reactions are observed according to the standards in Table 5 at 30 min (after the indentation disappears), 24 h, and 48 h after removing the test apparatus, and the results are recorded.

[0065] Result determination Interpretation of results from occlusive skin patch testing: If more than 5 out of 30 subjects have a suspected Grade 1 skin reaction, or more than 2 out of 30 subjects have a weakly positive Grade 2 skin reaction, or if any one subject has a Grade 3 or higher adverse skin reaction, the test substance is considered to have an adverse skin reaction on humans.

[0066] Table 5 Skin Reaction Scoring Criteria Rating levels Skin reaction 0 negative reaction 1 Suspicious reaction, only slight erythema 2 Weak positive reaction (erythema reaction): erythema, infiltration, edema, and papules may be present. 3 Strong positive reaction (herpes reaction): erythema, infiltration, edema, papules; the reaction may extend beyond the test area. 4 Extremely positive reaction (confluent herpes simplex reaction): marked erythema, severe infiltration, edema, confluent herpes simplex; reaction extends beyond the test area. After testing, the essence lotions 1-9, essence lotions 1'-5', and blank essence lotion provided by this invention all showed negative reactions after human patch testing, indicating that they are safe and non-irritating to human skin.

[0067] Test Example 4: Test on the effects of oil control, pore minimization, and skin firming on the human body. Subjects were selected based on the following inclusion criteria: oily or combination skin with a sebum secretion level ≥150 μg / cm³. 2 150 participants (healthy men and women aged 30-60 years) were divided into 15 groups (half male and half female) to test the product (Sebumeter assay). Each group used a sample of the test product (serum lotion 1-9 and serum lotion 1'-5', with a blank serum lotion as a control group). A sebum analyzer, VISIA 7, and a Cutometer MPA580 skin test probe were used to measure facial skin condition before product use (day 0), and on days 14 and 28 after product use. This allowed for a comprehensive evaluation of the product's oil control, pore-minimizing, and firming effects. Specific experimental parameters, equipment, and test indicators are shown in Table 6.

[0068] Table 6 Test Parameters and Equipment

[0069] The formulas for calculating changes in oil production, pore size, and skin firmness are as follows: Oil improvement rate = [(T0 sample group - T14 / T28 sample group) / T0 sample group × 100%] - [(T0 blank control group - T14 / 28 blank control group) / T0 blank application cases × 100%]; Pore ​​improvement rate = [(T0 sample group - T14 / T28 sample group) / T0 sample group × 100%] - [(T0 blank control group - T14 / 28 blank control group) / T0 blank control group × 100%]; Skin firmness improvement rate = [(T14 / T28 sample group - T0 sample group) / T0 sample group × 100%] - [(T14 / T28 blank control group - T0 blank control group) / T0 blank control group × 100%].

[0070] The results of the tests on the effects of oil control, pore shrinkage, and skin tightening on human skin are shown in Table 7. The data in the table show that on day 14 of using serum lotions 1-9, the subjects experienced significant improvements in facial oil secretion, pore volume, and skin tightness. Furthermore, the improvement effect continued to increase with prolonged use (e.g., day 28), indicating that the serum lotions provided by this invention have significant effects in oil control, pore shrinkage, and skin tightening. Compared to serum lotion 4, serum lotions 1'-5' showed significantly lower rates of oil improvement, pore volume improvement, and R2 value improvement. Moreover, the improvement in these effects was not significant with prolonged use. This indicates that adding a composition prepared from EGCG nanocrystals, medicinal blastocystis extract, Lactobacillus / pomegranate fermentation products, and cardamom seed ketone to the serum lotions can significantly improve their oil control, pore shrinkage, and skin tightening effects.

[0071] Table 7. Test Results of the Effects of Oil Control, Pore Minimizing, and Skin Firming on Human Body

[0072] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A composition for controlling oil, shrinking pores, and firming skin, characterized in that, The product comprises the following components in parts by weight: 0.01-0.5 parts EGCG nanocrystals, 0.01-4 parts medicinal blastocystis extract, 0.01-2.5 parts Lactobacillus / pomegranate fermentation product, and 0.001-0.1 parts cardamom seed ketone; wherein the EGCG nanocrystals are prepared by antisolvent crystallization of EGCG solution in a microchannel reactor using supercritical CO2 as the antisolvent.

2. The composition according to claim 1, characterized in that, It includes the following components in parts by weight: 0.05-0.3 parts EGCG nanocrystals, 0.1-3 parts medicinal blastocystis extract, 0.1-1 parts Lactobacillus / pomegranate fruit fermentation product, and 0.008-0.05 parts cardamom seed ketone.

3. The composition according to claim 1, characterized in that, It includes the following components in parts by weight: 0.08-0.1 parts EGCG nanocrystals, 0.5-1 parts medicinal blastocystis extract, 0.4-0.6 parts Lactobacillus / pomegranate fruit fermentation product, and 0.01-0.03 parts cardamom seed ketone.

4. The composition according to claim 1, characterized in that, The preparation method of the EGCG nanocrystals includes the following steps: S1. Dissolve EGCG in ethyl acetate, stir and sonicate under light-protected conditions, filter, collect the filtrate, and obtain an EGCG solution; S2. Using supercritical CO2 as the antisolvent, supercritical CO2 and EGCG solution were passed into a microchannel reactor for antisolvent crystallization to prepare EGCG nanocrystals.

5. The composition according to claim 4, characterized in that, The stirring in step S1 is magnetic stirring at a speed of 300-400 r / min for 15-25 min; the ultrasonic treatment is ultrasonication at an ultrasonic power of 90-100 W for 20-30 min.

6. The composition according to claim 4, characterized in that, The conditions for antisolvent crystallization in step S2 are as follows: at 35-37℃ and 10-12MPa pressure, the flow rate of supercritical CO2 is controlled at 33-42mL / min and the flow rate of EGCG solution is controlled at 1-2mL / min.

7. The composition according to claim 1, characterized in that, The preparation method of the Lactobacillus / pomegranate fruit fermentation product is as follows: After washing the mature pomegranate berries, add 5-10 times the weight of water, crush and pulp them to obtain pomegranate pulp, and sterilize it; inoculate the sterilized pomegranate pulp with Lactobacillus rhamnosus culture, and anaerobic ferment at 36-38℃ for 60-84h to obtain the fermentation product; centrifuge and filter the fermentation product to remove bacteria, obtain the fermentation broth, and freeze-dry it under vacuum to obtain the Lactobacillus / pomegranate fruit fermentation product.

8. The use of the composition according to any one of claims 1-7 in the preparation of cosmetics having the effects of oil control, pore shrinking and skin tightening.

9. The application as described in claim 8, characterized in that, The cosmetics include one of the following: toner, lotion, cream, mask, and spray.

10. An oil-controlling, pore-minimizing, and skin-firming essence lotion, characterized in that, The composition comprises the following components in weight percentages: 0.05%-0.5% thickener, 1%-10% humectant, 0.01%-0.3% pH adjuster, 1%-6% emulsifier, 5%-15% oil, 0.5%-3% preservative, 0.01%-0.3% chelating agent, 1%-5% of the composition according to any one of claims 1-7, and the balance being deionized water.

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