A Chinese medicine composition for treating acne and its preparation method and application

Abstract

The present invention provides a Chinese medicine composition for treating acne, a preparation method and an application thereof, and belongs to the technical field of Chinese medicine. The Chinese medicine composition comprises the following raw materials in parts by weight: 5 to 30 parts of Sophora flavescens, 5 to 30 parts of Salvia miltiorrhiza, 5 to 30 parts of Chrysanthemum indica, 5 to 30 parts of White Peony Root, and 1 to 10 parts of Licorice. The present invention provides a Chinese medicine composition for treating acne, a preparation method and an application thereof, the Chinese medicine composition has a small number of medicinal flavors, 5 selected Chinese medicines are properly compatible, the proportion is reasonable, and it has the effects of oil control, anti-inflammatory and antibacterial. The ingredients are simplified as much as possible while meeting the efficacy of treating acne, and the preparation method is simple, the ingredients are safe and have no toxic side effects, and can be used for a long time. The topical gel prepared by the Chinese medicine composition is accurately applied for 3 days, and the number and volume of acne are significantly reduced, and the product is mild and non-irritating, and has the effects of rapid acne removal and soothing.

Description

Technical Field

[0001] The invention belongs to the technical field of traditional Chinese medicines, and in particular relates to a traditional Chinese medicine composition for treating acne, and a preparation method and application thereof. Background Art

[0002] Acne is a chronic inflammatory skin disease that occurs in adolescence and mainly affects the pilosebaceous glands of the face. The causes of acne are relatively complex, and its pathogenesis mainly includes four factors: excessive secretion of lipids by sebaceous glands induced by hormones under genetic background, abnormal keratinization of pilosebaceous gland ducts, proliferation of follicular microorganisms such as Propionibacterium acnes, and inflammation and immune response, among which sebum production and inflammatory response are the main links. Acne generally manifests as polymorphic inflammatory skin lesions such as comedones, papules, pustules, nodules, cysts, etc., which have a significant impact on the patient's mood, daily activities, social activities and interpersonal relationships, and clinical diagnosis and treatment are more difficult.

[0003] At present, Western medicine mostly uses oral, topical drugs or physical therapy for treatment. Commonly used drugs include retinoic acid drugs, antibiotics, and hormones. Although Western medicine treatment has certain efficacy, there are still problems with contraindications and toxic side effects. For example, oral isotretinoin has toxic side effects such as teratogenicity and elevated blood lipids, and topical hormone chemicals may cause side effects such as chronic skin damage. Throughout the ages, the pathogenesis of acne is nothing more than "heat" and "depression". In the early stage of acne, "heat" is more important, while in the later stage, "depression" is more important. "Clearing heat" and "unblocking depression" are the basic treatments for acne. The connotation of the treatment method mainly includes clearing heat and detoxifying, dispersing wind and heat, clearing dampness and heat, promoting blood circulation and removing blood stasis, removing blood stasis and dispersing knots, and removing phlegm and unblocking collaterals. Traditional Chinese medicine uses oral Chinese medicine decoctions and Chinese medicine masks, acupuncture, bloodletting, acupoint thread embedding and other external treatments for treatment. Among them, external treatment with traditional Chinese medicine is a commonly used treatment method in clinical practice, but because the disease is self-limiting and easily causes fluctuations in the course of the disease, there are still great difficulties in external treatment with traditional Chinese medicine. Summary of the invention

[0004] In view of this, the object of the present invention is to provide a Chinese medicine composition for treating acne and a preparation method and application thereof. The Chinese medicine composition has good antibacterial, anti-inflammatory and oil-control effects and can treat acne efficiently and quickly.

[0005] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0006] The invention provides a traditional Chinese medicine composition for treating acne. The traditional Chinese medicine composition comprises the following raw materials in parts by weight: 5-30 parts of sophora flavescens, 5-30 parts of salvia miltiorrhiza, 5-30 parts of wild chrysanthemum, 5-30 parts of white peony root and 1-10 parts of liquorice.

[0007] Preferably, the Chinese medicine composition comprises the following raw materials in parts by weight: 10-25 parts of Sophora flavescens, 10-25 parts of Salvia miltiorrhiza, 10-25 parts of Chrysanthemum indici, 15-25 parts of White Peony Root, and 4-10 parts of Licorice.

[0008] The present invention also provides a method for preparing the above-mentioned Chinese medicine composition, comprising the following steps:

[0009] The raw materials are mixed with water, extracted, and the extract is concentrated to obtain fluid extract 1. Ethanol is added for alcohol precipitation, and the supernatant is taken and the ethanol is removed to obtain fluid extract 2.

[0010] Preferably, the mass ratio of the total mass of the raw materials to the mass of water is 1:6-10.

[0011] Preferably, the extraction temperature is 80-100° C., the extraction is performed 1-2 times, and the time for each extraction is 0.5-3 hours.

[0012] Preferably, the alcohol precipitation time is 10 to 15 hours.

[0013] Preferably, the mass volume ratio of the fluid extract 1 to ethanol is 1:1-3.

[0014] Preferably, the dosage form of the traditional Chinese medicine composition includes a gel, a patch, a solution or an ointment.

[0015] The present invention also provides an application of the above-mentioned Chinese medicine composition or the Chinese medicine composition obtained by the preparation method in preparing a drug for treating acne.

[0016] The present invention also provides a use of the above-mentioned Chinese medicine composition or the Chinese medicine composition obtained by the preparation method in the preparation of anti-inflammatory, oil-controlling or antibacterial drugs.

[0017] Compared with the prior art, the present invention has the following beneficial effects:

[0018] (1) The present invention provides a Chinese medicine composition for treating acne, and a preparation method and use thereof. The Chinese medicine composition has a small number of medicinal ingredients, and 5 kinds of Chinese medicines are selected to be properly compatible and reasonably proportioned. The ingredients are simplified as much as possible while satisfying the efficacy of treating acne. The preparation method is simple, the ingredients are safe, have no toxic side effects, and can be used for a long time.

[0019] (2) The Chinese medicine composition has no obvious cytological toxicity. It has no inhibition on the proliferation of HaCaT cells, human sebaceous gland cells (SZ95 cells), and mouse macrophages (RAW 264.7) within a certain concentration range, but can significantly inhibit lipid synthesis in human sebaceous gland cells, and has a good oil control effect. In addition, the Chinese medicine composition can inhibit the production of NO in the LPS-induced macrophage inflammation model, and has a good anti-inflammatory effect. The Chinese medicine composition also has an antibacterial effect and can inhibit the growth of Staphylococcus aureus and Propionibacterium acnes. The topical gel prepared with the Chinese medicine composition was accurately applied for 3 days, and the number and volume of acne were significantly reduced. The product is mild and non-irritating, and has the effects of quickly removing acne and soothing. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 This is a photo of the appearance of the Chinese medicine composition gel prepared in Example 6;

[0021] Figure 2 This is the TLC chart for identification of white peony root from different batches of fluid extract 2, from left to right: fluid extract 2-1, fluid extract 2-2, fluid extract 2-3, paeoniflorin;

[0022] Figure 3 This is the TLC chart for identification of Salvia miltiorrhiza of different batches of fluid extract 2, from left to right: fluid extract 2-1, fluid extract 2-2, fluid extract 2-3, and salvianolic acid B reference substance;

[0023] Figure 4 The TLC images of different batches of fluid extract 2 for identification of Sophora flavescens, from left to right: fluid extract 2-1, fluid extract 2-2, fluid extract 2-3, matrine;

[0024] Figure 5 This is the TLC chart for identification of wild chrysanthemum of different batches of fluid extract 2, from left to right: fluid extract 2-1, fluid extract 2-2, fluid extract 2-3, and monanthin;

[0025] Figure 6 TLC images of different batches of fluid extract 2 for identification of licorice, from left to right: fluid extract 2-1, fluid extract 2-2, fluid extract 2-3, licorice control medicinal material;

[0026] Figure 7 HPLC graphs of the determination of the content of lignoside in the lignoside reference substance and the Chinese medicine fluid extract 2, wherein a is the HPLC graph of lignoside in the lignoside reference substance, and A represents lignoside; b is the HPLC graph of lignoside in the Chinese medicine fluid extract 2, and A represents lignoside;

[0027] Figure 8 This is a representative photograph of subject 04;

[0028] Fig. 912 photographs of representative subjects. DETAILED DESCRIPTION

[0029] The invention provides a traditional Chinese medicine composition for treating acne. The traditional Chinese medicine composition comprises the following raw materials in parts by weight: 5-30 parts of sophora flavescens, 5-30 parts of salvia miltiorrhiza, 5-30 parts of wild chrysanthemum, 5-30 parts of white peony root and 1-10 parts of liquorice.

[0030] In the present invention, Sophora flavescens is cold and bitter, has the effects of clearing away heat and dampness, diuresis, dispelling wind, killing insects, etc., and can be used externally to treat skin itching and trichomonas vaginitis; the main components of Sophora flavescens are two categories of flavonoids and alkaloids, which have inhibitory effects on a variety of bacteria, have a wide range of physiological activities such as antibacterial, anti-inflammatory, antiviral, and immune regulation, and can reduce inflammatory reactions, thereby achieving the purpose of intervening in the formation of acne. The Sophora flavescens is preferably 10 to 25 parts, and the present invention has no special limitation on the source of Sophora flavescens, and commercially available products in this field can be used.

[0031] In the present invention, Danshen tastes bitter, slightly cold, enters the heart and liver meridians, has the effects of relieving pain, promoting blood circulation and removing blood stasis, clearing the heart and eliminating vexation, cooling blood and eliminating carbuncle, and plays the role of improving skin microcirculation, scavenging free radicals, anti-oxidation, anti-inflammatory, antibacterial, etc. The Danshen is preferably 10 to 25 parts, and the source of Danshen is not particularly limited in the present invention, and commercially available products in the field can be used.

[0032] In the present invention, wild chrysanthemum is slightly cold in nature, bitter in taste, and enters the liver and heart meridians, and has the effects of purging fire and calming the liver, clearing away heat and detoxifying. Wild chrysanthemum mainly contains flavonoids, volatile oils and various trace elements, among which montanol is an important flavonoid compound with significant antibacterial, anti-inflammatory and anti-aging effects. Wild chrysanthemum can clear away heat and detoxify, and has a certain effect in treating facial acne caused by heat toxins. The wild chrysanthemum is preferably 10 to 25 parts, and the present invention has no special limitation on the source of the wild chrysanthemum, and commercially available products in the art can be used.

[0033] In the present invention, white peony root is slightly cold in nature, bitter and sour in taste, and enters the liver and spleen meridians, and has the effects of nourishing blood and regulating menstruation, restraining yin and stopping sweating, softening the liver and relieving pain, and calming liver yang; the main components of white peony root include monoterpenes and their glycosides, triterpenes, flavonoids, tannins and polysaccharides, among which total glucosides of white peony root (TGP) is the main active ingredient of the traditional Chinese medicine white peony root, and has the effects of analgesia, anti-inflammatory, and liver protection. The white peony root is preferably 15 to 25 parts, and the present invention has no special limitation on the source of white peony root, and commercially available products in the field can be used.

[0034] In the present invention, licorice is sweet and neutral in nature, and its main effects are to tonify the spleen and replenish qi, clear away heat and detoxify, harmonize various medicines, eliminate phlegm and relieve cough, relieve acute pain, etc. Licorice chemical components include flavonoids, saponins, alkaloids, amino acids, coumarins, polysaccharides, etc., and have multiple pharmacological effects such as anti-oxidation, anti-inflammatory, immunomodulatory, anti-ulcer, anti-viral, anti-depressant, analgesic, liver protection, and anti-atherosclerosis. The licorice is preferably 4 to 10 parts, and the present invention has no special limitation on the source of licorice, and commercially available products in the field can be used.

[0035] The Chinese medicine composition of the present invention uses Sophora flavescens and Salvia miltiorrhiza as a pair of main drugs, which play the effects of clearing away heat and dampness, promoting blood circulation and removing blood stasis, reducing the permeability of capillaries in inflammatory tissues, reducing inflammatory exudation, and further reducing inflammatory damage to tissues; uses Wild Chrysanthemum as an assistant drug, which cooperates with the Sophora flavescens and Salvia miltiorrhiza drug pair to jointly play the role of treating acne; uses White Peony Root and Licorice Herb Pair as adjuvant drugs, which assist the main drugs Sophora flavescens and Salvia miltiorrhiza and the assistant drug Wild Chrysanthemum to jointly play the role of treating acne.

[0036] The present invention also provides a method for preparing the above-mentioned Chinese medicine composition, comprising the following steps:

[0037] The raw materials are mixed with water, extracted, and the extract is concentrated to obtain fluid extract 1. Ethanol is added for alcohol precipitation, and the supernatant is taken and the ethanol is removed to obtain fluid extract 2.

[0038] In the present invention, the mass ratio of the total mass of the raw materials to the mass of water is preferably 1:6 to 10. The present invention can better extract the effective ingredients such as paeoniflorin, salvianolic acid B, matrine and lignoside in the traditional Chinese medicine composition through water extraction, thereby achieving the purpose of treating acne.

[0039] In the present invention, in order to improve the extraction efficiency and prevent the active ingredients of the Chinese medicine composition of the present invention from being inactivated due to excessively high extraction temperature, the extraction temperature is preferably 80-100°C, more preferably 90-95°C. In order to shorten the extraction time and reduce the dissolution of ineffective ingredients, the number of extractions is preferably 1-2 times, more preferably 2 times. In order to ensure the efficacy of the Chinese medicine composition of the present invention and prevent the active ingredients from decreasing due to improper extraction time, the time for each extraction is preferably 0.5-3 hours, more preferably 1-2 hours.

[0040] In the present invention, ethanol is added for alcohol precipitation, the ethanol is 95% ethanol, the alcohol precipitation time is preferably 10 to 15 hours, more preferably 11 to 13 hours; the mass volume ratio of the fluid extract 1 to ethanol is preferably 1:1 to 3, which is equivalent to a volume percentage of ethanol of about 47 to 71%. The present invention uses the above-mentioned volume fraction of ethanol to precipitate the Chinese medicine composition to remove ineffective components, which can further improve the effect of treating acne.

[0041] In the present invention, the fluid extract 1 and the fluid extract 2 contain 0.5-1 g / mL of medicinal materials.

[0042] In the present invention, the dosage form of the Chinese medicine composition includes gel, patch, solution or ointment.

[0043] In the present invention, the preparation method of the gel of the traditional Chinese medicine composition comprises the following steps:

[0044] Take 0.5-1.5g of polymer material and 0.5-1g of ammonium acryloyldimethyltaurate / VP copolymer, add appropriate amount of water to swell them respectively, and obtain polymer material swelling liquid;

[0045] Take 220-40 mL of the above fluid extract, add it to the above polymer material swelling liquid, stir and mix, add a preservative, stir evenly, add a neutralizer to adjust the pH to 5.5-6.5, and finally add water to a total amount of 100 g, stir evenly, and the gel of the traditional Chinese medicine composition is obtained.

[0046] In the present invention, the polymer material is preferably carbopol, long-chain alkyl acrylate polymer; the carbopol is preferably carbomer 940 or cross-linked polyacrylic acid resin; the long-chain alkyl acrylate polymer is preferably acrylic acid (ester) C10-30 alkyl acrylate cross-linked polymer. The preservative is preferably one or more of phenoxyethanol, ethylhexylglycerin and p-hydroxyacetophenone. The neutralizer is preferably arginine and / or triethanolamine. In the present invention, in order to further improve the therapeutic effect of the above-mentioned topical gel, active ingredients such as 0.25% zinc glucose and 0.05% sodium hyaluronate can be added.

[0047] The present invention also provides an application of the above-mentioned Chinese medicine composition or the Chinese medicine composition obtained by the preparation method in preparing a drug for treating acne.

[0048] The present invention also provides a use of the above-mentioned Chinese medicine composition or the Chinese medicine composition obtained by the preparation method in the preparation of anti-inflammatory, oil-controlling or antibacterial drugs.

[0049] In the present invention, the Chinese medicine composition of the present invention can significantly inhibit Staphylococcus aureus and / or Propionibacterium acnes, and the antibacterial rate is more than 90%. The Chinese medicine composition can also inhibit the production of NO in the LPS-induced macrophage inflammation model, and has anti-inflammatory effects. At the same time, the Chinese medicine composition can significantly reduce the synthesis of neutral lipids, and has a good oil control effect.

[0050] In the present invention, the medicine comprises a Chinese medicine composition as an active ingredient and a pharmaceutically acceptable excipient. The medicine is administered by external application. The excipient preferably comprises one or more of a diluent, an adhesive, a wetting agent and a lubricant. The dosage of the Chinese medicine composition depends on many factors, such as the nature and severity of the disease to be prevented or treated, the patient's gender, age, weight, personality and individual response, administration route, administration frequency, and treatment purpose. Therefore, the daily dosage of the Chinese medicine composition of the present invention is 0.001 to 150 mg / kg body weight, preferably 0.01 to 100 mg / kg body weight, and more preferably 0.01 to 60 mg / kg body weight.

[0051] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0052] Example 1

[0053] A Chinese medicine composition for treating acne, comprising the following raw materials in parts by weight:

[0054] 20 parts of Sophora flavescens, 20 parts of Salvia miltiorrhiza, 20 parts of Flos Chrysanthemi Indici, 20 parts of the White Peony Roots, 10 parts of Radix Glycyrrhizae.

[0055] Take the weight portion of the Chinese medicine composition and put it into an extraction tank, add 8 times the amount of water, extract for 2 hours at 95° C. for the first time, extract for 1 hour at 95° C. for the second time, combine the two extracts, transfer to a concentration tank, concentrate under reduced pressure to obtain a fluid extract 1 (containing 1 g / mL of the medicinal material), cool to room temperature, add 195 parts of ethanol to make the alcohol content reach 65%, and precipitate with alcohol for 12 hours. Take the supernatant, recycle the ethanol under reduced pressure until there is no alcohol taste, and obtain a fluid extract 2 (containing 1 g / mL of the medicinal material), which is reserved.

[0056] Example 2

[0057] A Chinese medicine composition for treating acne, comprising the following raw materials in parts by weight:

[0058] 5 parts of Sophora flavescens, 30 parts of Salvia miltiorrhiza, 25 parts of Flos Chrysanthemi Indici, 30 parts of the White Peony Roots, 5 parts of Radix Glycyrrhizae.

[0059] Take the weight portion of the Chinese medicine composition and put it into an extraction tank, add 7 times the amount of water, extract for 1 hour at 80° C. for the first time, and extract for 1 hour at 90° C. for the second time, combine the two extracts, transfer to a concentration tank, and concentrate under reduced pressure to obtain a fluid extract 1 (containing 1 g / mL of the medicinal material), cool to room temperature, add 265 parts of ethanol to make the alcohol content reach 70%, and precipitate with alcohol for 10 hours. Take the supernatant, recycle the ethanol under reduced pressure until there is no alcohol taste, and obtain a fluid extract 2 (containing 1 g / mL of the medicinal material), which is reserved.

[0060] Example 3

[0061] A Chinese medicine composition for treating acne, comprising the following raw materials in parts by weight:

[0062] 30 parts of Sophora flavescens, 5 parts of Salvia miltiorrhiza, 20 parts of Flos Chrysanthemi Indici, 5 parts of White Peony Root, and 1 part of Licorice.

[0063] Take the weight portion of the Chinese medicine composition and put it into an extraction tank, add 6 times the amount of water, extract at 100° C. for 3 hours to obtain an extract, transfer it to a concentration tank, concentrate under reduced pressure to obtain a fluid extract 1 (containing 1 g / mL of the medicinal material), cool it to room temperature, add 105 parts of ethanol to make the alcohol content reach 60%, and precipitate it with alcohol for 15 hours. Take the supernatant, reduce the pressure to recover ethanol until there is no alcohol taste, and obtain a fluid extract 2 (containing 1 g / mL of the medicinal material) for standby use.

[0064] Example 4

[0065] A Chinese medicine composition for treating acne, comprising the following raw materials in parts by weight:

[0066] 25 parts of Sophora flavescens, 15 parts of Salvia miltiorrhiza, 5 parts of Flos Chrysanthemi Indici, 15 parts of the White Peony Roots, 7 parts of Radix Glycyrrhizae.

[0067] Take the weight portion of the Chinese medicine composition and put it into an extraction tank, add 8 times the amount of water, extract for 2 hours at 90° C. for the first time, extract for 1 hour at 90° C. for the second time, combine the two extracts, transfer to a concentration tank, concentrate under reduced pressure to obtain a fluid extract 1 (containing 1 g / mL of the medicinal material), cool to room temperature, add 145 parts of ethanol to make the alcohol content reach 65%, and precipitate with alcohol for 13 hours. Take the supernatant, recover the ethanol under reduced pressure until there is no alcohol taste, and obtain a fluid extract 2 (containing 1 g / mL of the medicinal material), which is reserved.

[0068] Example 5

[0069] A method for preparing a gel of a traditional Chinese medicine composition, comprising the following steps:

[0070] Take 0.8g of acrylic acid (ester) C10-30 alkyl acrylate cross-linked polymer and 0.5g of ammonium acryloyldimethyltaurate / VP copolymer, add appropriate amount of water to swell, and obtain a polymer material swelling solution; take 20mL of the Chinese medicine fluid extract 2 (1g medicinal material / mL) prepared in Example 1, add it to the above polymer material swelling solution, stir and mix, add 0.25g of zinc gluconate, 1g of polyglycerol-31g, 0.05g of sodium hyaluronate, 0.5g of phenoxyethanol, 0.1g of ethylhexylglycerol and 0.1g of parahydroxyacetophenone, stir evenly, add 1.5g of arginine to adjust the pH to 5.5-6.5, and finally add water to the total amount of 100g, stir evenly, and obtain.

[0071] Example 6

[0072] Formula of Chinese medicine composition gel:

[0073]

[0074] Preparation method: Take the prescribed amount of Carbomer 940 and swell it with an appropriate amount of pure water overnight, dissolve zinc gluconate, potassium azelayl diglycinate, and sodium hyaluronate in the remaining amount of pure water, stir with a blender until the raw materials are dissolved, add Chinese medicine fluid extract 2 and phenoxyethanol and continue to stir evenly, mix with the swollen Carbomer 940, stir evenly, add triethanolamine to make it gel. Among them, the appearance photo of the gel is shown in Figure 1 .

[0075] Example 7

[0076] Formula of Chinese medicine composition gel:

[0077]

[0078] Test Example 1

[0079] Example 1 Preparation of Chinese medicine composition fluid extract 2 Quality standard

[0080] (1) Identification of white peony root in fluid extract 2: Take 20.5 mL of fluid extract, add 10 mL of ethanol, shake for 5 minutes, filter, evaporate the filtrate, add 1 mL of ethanol to dissolve the residue, and use it as the test solution. Take another reference substance of paeoniflorin, add ethanol to make a solution containing 1 mg per 1 mL, and use it as the reference substance solution. According to the thin layer chromatography method (General Rule 0502), take 10 μL of each of the above two solutions, spot them on the same silica gel G thin layer plate, use chloroform-ethyl acetate-methanol-formic acid (40:5:10:0.2) as the developing solvent, develop, take out, dry, spray with 5% vanillin sulfuric acid solution, and heat until the spots are clear.

[0081] Figure 2 The results show that the Chinese medicine composition fluid extract 2 of the present invention contains paeoniflorin.

[0082] (2) Identification of Salvia miltiorrhiza in fluid extract 2: Take 21 mL of fluid extract, heat and evaporate to dryness, add 2 mL of ethanol to dissolve, and use it as the test solution. Take 1 g of Salvia miltiorrhiza reference medicinal material, add 5 mL of ethanol, ultrasonicate for 15 minutes, centrifuge, and take the supernatant as the reference medicinal material solution. Take Salvianolic acid B reference substance, add ethanol to make a solution containing 1.5 mg per 1 mL, and use it as the reference substance solution. According to the thin layer chromatography method (General Rule 0502), take 3 μL of each of the above three solutions and spot them on the same silica gel G thin layer plate to make strips. Use chloroform-toluene-ethyl acetate-methanol-formic acid (6:4:8:1:4) as the developing solvent, develop to about 4 cm, take out, dry, and then use petroleum ether (60-90℃)-ethyl acetate (4:1) as the developing solvent, develop to about 8 cm, take out, dry, and examine under sunlight and ultraviolet light (365nm).

[0083] Figure 3 The results show that the Chinese medicine composition fluid extract 2 of the present invention contains salvianolic acid B.

[0084] (3) Identification of Sophora flavescens in fluid extract 2: Take 210 mL of fluid extract, add 1 mL of concentrated ammonia test solution, add 20 mL of chloroform, ultrasonicate in a water bath for 45 min, filter and evaporate to dryness, add 2 mL of methanol to dissolve the residue, which is the test solution; take matrine reference substance and add ethanol to make a solution with a concentration of 0.2 mg / mL as the reference solution. According to the thin layer chromatography method (General Rule 0502), take 4 μL of each of the above solutions and spot it on the same silica gel G thin layer plate, use the lower layer solution of chloroform: acetone: ammonia water (10:2:1) as the developing agent, develop, take out, dry, and spray with potassium bismuth iodide test solution.

[0085] Figure 4 The results show that the Chinese medicine composition fluid extract 2 of the present invention contains matrine.

[0086] (4) Identification of wild chrysanthemum in fluid extract 2: Take 21 mL of fluid extract, add 15 mL of methanol, ultrasonicate in a water bath for 45 min, filter, heat and concentrate to 5 mL, as the test solution. Take another reference substance of monanthin, add methanol to make a solution with a concentration of 0.2 mg / mL, as the reference solution. According to the thin layer chromatography method (General Rule 0502), take 5 μL of each of the above solutions and spot them on the same polyamide film, use the lower layer solution of ethyl acetate: butanone: chloroform: formic acid: water (15:15:6:4:1) as the developing agent, develop, take out, dry, spray with aluminum chloride test solution, blow dry with hot air, and inspect under ultraviolet light (365 nm).

[0087] Figure 5 The results show that the Chinese medicine composition fluid extract 2 of the present invention contains limonene glycoside.

[0088] (5) Identification of Licorice in Fluid Extract 2: Take 21 mL of fluid extract, add 40 mL of ether, heat and reflux for 1 hour, filter, discard the ether solution, add 30 mL of methanol to the drug residue, heat and reflux for 1 hour, filter, evaporate the filtrate to dryness, add 40 mL of water to the residue to dissolve, extract with n-butanol 3 times, 20 mL each time, combine the n-butanol solution, wash with water 3 times, discard the water solution, evaporate the n-butanol solution to dryness, add 5 mL of methanol to the residue to dissolve, and use it as the test solution. Take 1 g of licorice control medicinal material and prepare the control medicinal material solution in the same way. According to the thin layer chromatography method (general column 0502), 1-2 μL of each of the above two solutions were taken and spotted on the same silica gel G thin layer plate, and ethyl acetate-formic acid-glacial acetic acid-water (15:1:1:2) was used as the developing agent. After development, the plate was taken out and dried, and sprayed with 10% ethanolic sulfuric acid solution. The plate was heated at 105°C until the spots were clearly colored and examined under ultraviolet light (365nm).

[0089] Figure 6The results show that the Chinese medicine composition fluid extract 2 of the present invention contains licorice components.

[0090] (6) HPLC determination method of the content of monanthin in fluid extract 2:

[0091] S1. Chromatographic conditions and system suitability test: Octadecylsilane bonded silica gel was used as filler (5 μm, 250 mm × 4.6 mm, Agilent, USA); methanol-0.1% phosphoric acid solution (58:42) was used as mobile phase; detection wavelength was 334 nm, column temperature was 30 °C, flow rate was 1 mL / min. The theoretical plate number calculated based on the limonene peak should be no less than 3000.

[0092] S2. Preparation of reference solution: Take an appropriate amount of linalool reference substance, weigh it accurately, add methanol and ultrasonically dissolve it to make a solution containing 8 μg per 1 mL to obtain the reference solution.

[0093] S3. Preparation of test solution: Take 21 mL of the product fluid extract and put it into a conical flask, add 19 mL of methanol, weigh it, sonicate in a water bath for 15 min, cool it, add methanol to make up the weight, shake it well, filter it through a 0.45 μm filter membrane, and take the filtrate to obtain the test solution.

[0094] Determination method: Accurately pipette 10 μL of reference solution and test solution respectively, inject into liquid chromatograph, and determine. The chromatogram of the determined mongoside is shown in Figure 7 .

[0095] The above-mentioned HPLC detection method was used to determine the content of montanaglycoside in fluid extract 2. The content of montanaglycoside (C montanaglycoside) per gram of crude drug (1 g of medicinal material / mL) in three batches of fluid extract 2 was 1.3477 g / mL. 28 H 32 O 14 ) were 596.58, 578.8 and 625.26 μg respectively.

[0096] 4. In vitro cytological efficacy study

[0097] In vitro studies on the soothing, oil-control, anti-inflammatory and antibacterial effects of the product were conducted using human immortalized keratinocyte cell line HaCaT cells, human sebaceous gland cells SZ95, mouse monocyte-macrophage cell line RAW264.7 cells, Staphylococcus aureus and Propionibacterium acnes.

[0098] (1) Cytological toxicity evaluation of the Chinese medicine fluid extract 2 prepared in Example 1

[0099] Main reagents and materials: DMEM culture medium, fetal bovine serum (FBS), penicillin + streptomycin double antibody, purchased from Gibco; T25 cell culture flasks, 96-well plates, purchased from Corning; cell counting kit (cck)-8, purchased from Dojindo Molecular Technologies; phosphate buffered saline (PBS, 1X), purchased from Biyuntian; Chinese medicine dry extract powder (each gram of dry extract powder is equivalent to 8.3 grams of crude drug, which is obtained by further vacuum drying or freeze-drying the aforementioned fluid extract 2).

[0100] Cell lines and cell culture: The cell lines used in this study were human immortalized keratinocytes HaCaT (Shanghai Traditional Chinese Medicine External Preparation Innovation Engineering Technology Research Center). HaCaT cells were cultured in a cell culture incubator (5% CO2, 37°C) using a medium containing DMEM+10% FBS+1% double antibody.

[0101] Determination of HaCaT cell proliferation rate: Use the cck-8 kit to test cell viability. 10,000 HaCaT cells were inoculated in each well of a 96-well plate and grown under culture conditions for 24 hours. Subsequently, the supernatant was aspirated. Traditional Chinese medicine extracts of different dilution multiples (25μg / mL, 50μg / mL, 100μg / mL, 200μg / mL, 400μg / mL) were added to each well and incubated for 24 hours, with 5 replicates in each group, and the same amount of PBS was added to the control group. After incubation, the supernatant was discarded, 100uL of DMEM containing 10% CCK8 was added, and the cell OD was measured after incubation in a cell culture incubator for 30 minutes. 450 .

[0102] Cell viability (%) = [OD 450 (Test)-OD 450 (PBS well)] / [OD 450 (Positive)-OD 450 (PBS well)] Experimental results and conclusions:

[0103] Table 1 Effects of different concentrations of Chinese medicine fluid extract 2 on the proliferation ability of HaCaT cells

[0104]

[0105] The results in Table 1 show that when the concentration of the Chinese herbal extract does not exceed 400 μg / mL, it has no significant effect on the proliferation of HaCaT cells, indicating that it has no cytotoxicity.

[0106] (2) Study on the oil control effect of Chinese herbal fluid extract 2

[0107] Main reagents: DMEM culture medium, fetal bovine serum (FBS), penicillin + streptomycin, and double antibody were purchased from Gibco; T25 cell culture flasks and 96-well plates were purchased from Corning; cell counting kit (cck)-8 was purchased from Dojindo Molecular Technologies, and phosphate buffered saline (PBS, 1X) was purchased from Biyuntian; Chinese medicine dry extract powder (each gram of dry extract powder is equivalent to 8.3 g of crude drug, which is obtained by further vacuum drying or freeze-drying the aforementioned fluid extract 2).

[0108] Cell line and cell culture: Human sebaceous gland cells SZ95 (provided by Shanghai Traditional Chinese Medicine External Preparation Innovation Engineering Technology Research Center). SZ95 was cultured in a cell culture incubator (5% CO2, 37°C) using a medium containing DMEM+10% FBS+1% double antibody.

[0109] Effect of Chinese herbal extract on SZ95 cell proliferation: SZ95 cells were inoculated in 96-well plates at 10,000 cells per well, with 3 replicates per group. Grouping: 3 extract concentrations were set, namely 50μg / mL, 100μg / mL, and 200μg / mL. After inoculating the cells, they were allowed to adhere to the wall overnight for 24 hours, and then different concentrations of Chinese herbal extract or DMEM medium were added and incubated for 24 hours. The supernatant was discarded, 10% CCK8100uL was added, and the cells were incubated in a cell culture incubator for 30 minutes. The absorbance of 450 was detected using a multifunctional microplate reader. Cell growth rate = (OD value of experimental well - OD value of PBS well) / (OD value of blank well - OD value of PBS well).

[0110] Effect of Chinese herbal extract on neutral lipid synthesis in SZ95 cells: Cells were inoculated in a black-bottomed 96-well plate (10,000 cells per well). The next day, different concentrations of Chinese herbal extract were added and incubated for 24 hours, then the supernatant was discarded. The cells were washed with PBS for 2 hours and stained with 10 μg / mL Nile red diluted in PBS. The cells were incubated at 37°C in the dark for 10 minutes, and the fluorescence intensity of Nile red was detected using a multifunctional microplate reader with an excitation wavelength of 485 nm / emission wavelength.

[0111] Experimental results and conclusions:

[0112] Table 2 Effects of different concentrations of Chinese medicine fluid extract 2 on the proliferation ability of SZ95 cells

[0113]

[0114] The results in Table 2 show that the Chinese herbal extract has no inhibitory effect on the proliferation of SZ95 cells, that is, it has no cytotoxicity.

[0115] Table 3 Effects of Chinese medicine fluid extract 2 on neutral lipid synthesis in SZ95 cells

[0116]

[0117] Conclusion: The Chinese herbal extract at a concentration of 200 μg / mL can significantly reduce the fluorescence intensity of Nile red staining, that is, reduce the synthesis of neutral lipids, indicating that it has a good oil control effect.

[0118] (3) Study on the anti-inflammatory efficacy of Chinese medicine fluid extract 2

[0119] Experimental cells: Mouse mononuclear macrophage RAW 264.7 cell line.

[0120] Test drugs and reagents: Chinese medicine dry extract powder (each gram of dry extract powder is equivalent to 8.3g of crude drug); lipopolysaccharide LPS, DMEM high glucose culture medium, fetal bovine serum, CCK8 detection kit, nitric oxide (NO) detection kit.

[0121] Instruments and equipment: biological safety cabinet, cell constant temperature incubator, inverted microscope, fully automatic microplate reader, centrifuge, water bath, -80 degree refrigerator, -20 degree refrigerator, 4 degree refrigerator, 1 / 10,000 balance, 96-well plate, 12-well plate, disposable spatula, disposable pipette, guns and gun tips of different specifications such as 1000μL, 200μL, 10μL, 50mL, 15mL and 2mL centrifuge tubes.

[0122] RAW264.7 cell culture: Mouse mononuclear macrophage RAW 264.7 cell line was cultured in DMEM high glucose medium containing 10% FBS at 37°C and 5% CO2 constant temperature incubator. The cells grew logarithmically, and the medium was replaced every 24 hours. The cells were passaged when they grew to 80% to 90%. After the cells were passaged to the third generation and the state was stable, they were used for experiments.

[0123] Drug preparation: dilute the Chinese medicine dry extract powder to 400μg / mL, 200μg / mL, 100μg / mL, and 50μg / mL.

[0124] CCK8 assay to detect the activity of RAW 264.7 cells: To investigate the effect of Chinese herbal extract on the activity of mouse mononuclear macrophage RAW 264.7 cells, macrophages in the logarithmic growth phase were scraped with a disposable spatula and blown with a pipette until they were dispersed into single cells. The corresponding number of cells was counted and the corresponding culture medium was added at 4×10 4 / mL density was seeded into 96-well plates, 100uL per well, to make it adhere to the wall. The experiment set up a blank control group, and the concentrations of Chinese medicine fluid extract 2 were diluted to 50μg / mL, 100μg / mL, 200μg / mL, and 400μg / mL, with 5 replicates in each group. The blank control group added the corresponding volume of cell culture fluid and continued to incubate for 24h. Discard the upper culture medium of the 96-well plate, add 100μL of 10% CCK8 serum-free DMEM solution, incubate in a cell culture incubator at 37℃ and 5% CO2 for 30min, measure the OD450 value with an enzyme marker, and calculate the cell survival rate. Cell survival rate (%) = (OD value of Chinese medicine group-OD value of PBS well) / (OD value of blank control group-OD value of PBS well) × 100%.

[0125] Nitrate reductase method to detect NO content: Take the cells under "2.1" and add 20×10 4 100 μL of 10μg / mL LPS was added to the drug group and the model control group, and the blank group was added with complete culture medium. The culture was continued for 24 hours, and the supernatant was collected and tested in a 96-well plate. The specific steps were operated according to the instructions of the NO kit. The absorbance was measured at 540nm, and the NO content and inhibition rate were calculated. Formula: The NO content was substituted into the formula for calculation. Formula: NO inhibition rate (%) = (average NO content in the model control group - average NO content in the drug group) / average NO content in the model control group × 100%.

[0126] Experimental results and conclusions:

[0127] Table 4 Effects of different concentrations of Chinese herbal extracts on the proliferation of RAW264.7 cells

[0128]

[0129]

[0130] The results in Table 4 show that the Chinese medicine fluid extract 2 has no inhibitory effect on the growth of macrophages in the range of 50 μg / mL to 400 μg / mL, indicating that it has no cytotoxicity.

[0131] Preparation of standard curve for detecting NO content by nitrate reductase method: According to the operating steps of the kit, a linear relationship between NO concentration and OD value was established. The results are shown in the figure below. The linear equation is: y=0.0066x+0.1125 (r=1).

[0132] Table 5 Effects of different concentrations of Chinese medicine fluid extract 2 on NO release in RAW 264.7 cells

[0133]

[0134] The results in Table 5 show that Chinese medicine fluid extract 2 can inhibit the production of NO in the LPS-induced macrophage inflammation model in a dose-dependent manner, indicating that it has a good anti-inflammatory effect.

[0135] (4) Evaluation of the antibacterial efficacy of Chinese herbal extracts

[0136] Strains: The strains used in this study: Staphylococcus aureus ATCC 6538 originated from Shanghai Dermatology Hospital, and Propionibacterium acnes ATCC 6919 was purchased from Beina Biotechnology Co., Ltd. (Cat. No.: BNCC336649).

[0137] Main reagents: nutrient broth (NB), soybean casein agar (TSA), brain heart infusion (BHI) broth, brain heart infusion (BHI) agar, skim milk culture tube, anaerobic generation bag, anaerobic culture bag, oxygen indicator were all purchased from Haibo Biotechnology; 96-well plate was purchased from Corning; Chinese medicine extract was homemade.

[0138] Main instruments: Spectramax M2 multi-mode microplate reader (Molecular Devices, USA), MaxQ TM 6000 Constant Temperature Shaker Incubator, Thermo Scientific TM 1300 biological safety cabinet.

[0139] Bacterial culture: During the experiment, strains of Propionibacterium acnes SC 5314 and Staphylococcus aureus ATCC 6538 were stored in skim milk culture tubes at -80°C for a long term.

[0140] Staphylococcus aureus: Before culturing, take out the frozen strain, streak it onto the TSA agar plate with an inoculation loop after freeze-thawing, and culture it at 37°C for 24-48h until there are obvious colonies as the working plate. The working plate is replaced every 20 days to ensure the activity of the colonies and stored at 4°C. To obtain the Staphylococcus aureus culture after the logarithmic growth period, pick a small round colony on the working plate each time, place it in 1mL NB liquid culture medium, shake the bacteria at a speed of 250rpm and a temperature of 37°C overnight, and harvest the strain for the experiment the next day.

[0141] Propionibacterium acnes: Propionibacterium acnes is an anaerobic bacterium and needs to be cultured in an anaerobic environment. An opened anaerobic bag is placed in the anaerobic culture bag to create an anaerobic environment, and an anaerobic indicator is used to observe the oxygen content in the culture bag. Before culture, the frozen strain is taken out, and after freezing and thawing, it is streaked onto the BHI agar plate using an inoculation loop, and placed in an anaerobic environment at 37°C for 48 to 72 hours until there are obvious colonies as the working plate. The working plate is replaced every 20 days to ensure the activity of the colony and stored at 4°C. In order to obtain the Propionibacterium acnes bacterial liquid after the logarithmic growth period, a small circular colony on the working plate is picked each time, placed in 1mL BHI liquid culture medium, and shaken overnight in an anaerobic environment at a speed of 250rpm and a temperature of 37°C. The strain is harvested the next day for experiment.

[0142] Determination of the antibacterial rate of Chinese medicine fluid extract 2 against Staphylococcus aureus and Propionibacterium acnes: The micro-double dilution method is used to test the antibacterial rate of Chinese medicine fluid extract 2 solution (containing 1g / mL of medicinal materials) against Staphylococcus aureus and Propionibacterium acnes. The specific steps are as follows:

[0143] (1) Add 100 μL of culture medium (NB-Staphylococcus aureus; BHI-Propionibacterium acnes) to each well of a 96-well plate;

[0144] (2) Add 100 μL of the prepared Chinese medicine fluid extract 2 solution to the first row, and then dilute the drug twice. That is, after adding the Chinese medicine extract solution to the first well, use a pipette to mix it thoroughly, then pipette 100 μL into the second well and mix it thoroughly, and repeat this process until the last well. Pipet 100 μL from the last well and discard it. Repeat this process 3 times for each drug concentration. At this point, each test well should contain 100 μL of solution.

[0145] (3) Add 100 μL of diluted Staphylococcus aureus or Propionibacterium acnes bacterial solution to each well (the concentration of the diluted bacterial solution is 2×10 6 CFU / mL), and the final concentration of the bacterial solution added to each well was 1×10 6 CFU / mL, the final dilution factor of the Chinese medicine extract solution in the first well was 4 times;

[0146] (4) Set up a negative control on the same plate (containing drugs and culture medium at different dilutions to remove drug background OD 630 ) and positive control (100 μL culture medium + 100 μL bacterial solution, the final concentration of bacterial solution was 1×10 6 CFU / mL)

[0147] (5) After incubation at 50 rpm for 24 hours, the OD630 of each well was measured using a microplate reader to calculate the bacterial activity.

[0148] Inhibition rate (%) = 100-[OD 630(Test)-OD 630 (negative)] / [OD 630 (Positive)-OD 630 (Negative)]

[0149] Experimental results and conclusions:

[0150] Table 6 Inhibition rate of Chinese medicine fluid extract 2 against Staphylococcus aureus at different dilution times (4 to 256 times)

[0151] Dilution multiple 4 times 8 times 16 times 32 times 64 times 128 times 256 times Positive OD1 0.408 0.288 0.215 0.273 0.314 0.35 0.421 0.525 OD2 0.404 0.27 0.211 0.271 0.307 0.327 0.435 0.515 OD3 0.391 0.266 0.208 0.227 0.304 0.361 0.398 0.498 Negative control 0.402 0.253 0.187 0.153 0.13 0.139 0.128 0.122 Antibacterial rate (%) 100.26％ 94.45％ 93.77％ 73.38％ 54.35％ 47.01％ 25.77％ SD(%) 1.86％ 2.45％ 0.73％ 5.43％ 1.07％ 3.63％ 3.90％

[0152] Note: Yang represents the positive control group, SD represents the standard deviation.

[0153] As shown in Table 6, the antibacterial rates of the Chinese medicine fluid extract 2 diluted 4 to 16 times against Staphylococcus aureus were all above 90%, and the antibacterial rate was still 25.77% at a 256-fold dilution concentration.

[0154] Table 7 Inhibition rate of Chinese herbal extracts at different dilution times (4-256 times) against Propionibacterium acnes

[0155] Dilution multiple 4 times 8 times 16 times 32 times 64 times 128 times 256 times Positive OD1 0.188 0.154 0.152 0.275 0.411 0.42 0.435 0.408 OD2 0.204 0.17 0.121 0.261 0.407 0.453 0.422 0.415 OD3 0.196 0.169 0.144 0.257 0.392 0.467 0.464 0.438 Negative control 0.192 0.158 0.123 0.104 0.092 0.091 0.083 0.06 Antibacterial rate (%) 98.89％ 98.24％ 95.56％ 55.50％ 13.60％ 1.30％ 0.83％ SD(%) 1.81％ 2.03％ 3.65％ 2.14％ 2.27％ 5.47％ 4.87％

[0156] Note: Yang represents the positive control group, SD represents the standard deviation.

[0157] As shown in Table 7, the antibacterial rates of the Chinese medicine fluid extract 2 diluted 4 to 16 times against Propionibacterium acnes were all above 90%, the antibacterial rate of the Chinese medicine fluid extract 2 diluted 64 times could still inhibit about 13.60% of Propionibacterium acnes, and the antibacterial effect was almost non-existent at a dilution factor of 128 to 256 (inhibition rate <10%).

[0158] The above results show that Chinese medicine fluid extract 2 can inhibit the growth of Staphylococcus aureus and Propionibacterium acnes, indicating that it has an antibacterial effect, and the more concentrated the drug solution, the better the inhibitory effect.

[0159] Test Example 2

[0160] Experimental materials: the Chinese medicine composition gel prepared in Example 5.

[0161] Human efficacy test of the topical gel prepared in Example 5 for treating acne:

[0162] Human efficacy test basis:

[0163] (1) T / GDCDC 023-2022 "Test Methods for Anti-Acne Efficacy of Cosmetics"

[0164] (2) T / GDCDC 021-2022 "Test Method for Soothing Effect of Cosmetics"

[0165] (3) Liu Weiyi, Zhou Lin, Zhao Hua. Evaluation of cosmetic efficacy (ⅩⅢ) - Consumer use test. Daily Chemical Industry, 2021; 51(06): 485-490

[0166] Specific test plan:

[0167] The subjects themselves were used for before and after control. After cleaning their faces, the subjects sat quietly for 30 minutes in a standard environment (temperature 20℃~22℃, humidity 40%~60%), Visia image acquisition + IPP analysis of the red zone, 3D skin imaging analyzer Antera-3D analyzed the acne volume / redness average value, recorded as D0, the subjects took the samples back for trial use, and revisited the test on the 1st and 3rd days, recorded as D1 and D3. When using, the subjects took an appropriate amount of gel on a cotton swab, accurately applied to the acne and its surrounding areas, taking care to avoid the skin around the eyes, and applied once every 4 hours. Use 4 times a day.

[0168] Inclusion criteria:

[0169] 1. Healthy Chinese women or men, aged between 18 and 40 years old;

[0170] 2. Those with acne in the test area;

[0171] 3. The subjects do not have severe chronic wasting diseases (asthma, diabetes, etc.);

[0172] 4. The subjects have no skin lesions, inflammation, eczema, etc.;

[0173] 5. The subjects have no hair, skin lesions or other factors that may affect the measurement at the test site;

[0174] 6. The subjects can complete the trial as required, read and understand all the contents of the informed consent form, and voluntarily sign the informed consent form.

[0175] Exclusion criteria:

[0176] 1. Those who have used antihistamines in the past week or immunosuppressants in the past month;

[0177] 2. Those who have used any anti-inflammatory drugs in the tested area within the past two months;

[0178] 3. Subjects with inflammatory skin diseases that have not been clinically cured;

[0179] 4. Patients with insulin-dependent diabetes;

[0180] 5. Patients who are receiving treatment for asthma or other chronic respiratory diseases;

[0181] 6. Those who have received anti-cancer chemotherapy in the past 6 months;

[0182] 7. Patients with immunodeficiency or autoimmune diseases;

[0183] 8. Lactating or pregnant women;

[0184] 9. Patients who have undergone bilateral mastectomy and bilateral axillary lymph node removal;

[0185] 10. The judges whose test results may be affected by scars, pigments, atrophy, port wine stains or other blemishes on the skin to be tested;

[0186] 11. Participants in other clinical trials;

[0187] 12. People with highly sensitive constitution;

[0188] 13. Non-voluntary participants or those who cannot complete the required content according to the test requirements;

[0189] Exit criteria:

[0190] 1. The subject is lost to follow-up or actively requests to withdraw;

[0191] 2. Poor compliance, failure to use the product on time and in the prescribed amount, and failure to make return visits as required;

[0192] 3. Violation of the research protocol;

[0193] 4. Serious adverse reactions occur during the entire testing period.

[0194] Criteria:

[0195] If any of the parameters including the average redness value and acne volume of the test area show significant improvement before and after product use, it indicates that the gel has the effect of treating acne; if the average optical density of skin erythema in the test area shows significant improvement, it indicates that the gel has a soothing effect; the questionnaire options use a 5-point method, and the test results require that the percentage of subjects agreeing is greater than 50% to support the relevant claims.

[0196] Specific test results:

[0197] A total of 32 subjects were recruited, and 30 of them finally completed the test, including 21 females and 9 males with an average age of 30.07±6.14 years, which met the voluntary inclusion criteria.

[0198] In accordance with the "Technical Specifications for Safety of Cosmetics" (2015 edition), each subject used the product once, and the adverse reaction record results showed no adverse reactions occurred.

[0199] 1. Average redness

[0200] Table 8 Redness average value test results 1

[0201] Time quantity Mean Standard Deviation Minimum Median Maximum Before use (D0) 30 32.89 9.36 21.10 34.50 53.10 After use (D1) 30 31.36 9.25 18.60 31.60 53.40 After use (D3) 30 28.72 7.60 18.00 27.65 46.40

[0202] Table 9 Redness average value test results 2

[0203]

[0204]

[0205] Average redness value: represents the average redness value of the skin. The larger the value, the higher the redness of the skin.

[0206] Table 10 Redness average value test results 3

[0207]

[0208] “ns” indicates no statistical difference, P>0.05; “*” indicates significant difference, 0.01≤P<0.05, noted as “*”; 0.001≤P<0.01, noted as “**”; p<0.001, noted as: “***”.

[0209] After continuous use of the gel for 3 days, the average redness of the subjects decreased by 4.65% compared with the baseline value on D1, and decreased significantly (P < 0.001) by 12.68% on D3.

[0210] 2. Acne volume

[0211] Table 11 Acne volume detection results 1

[0212] Time quantity Mean Standard Deviation Minimum Median Maximum Before use (D0) 30 0.9634 0.9690 0.1280 0.7095 4.8100 After use (D1) 30 0.9049 0.9095 0.0242 0.5980 4.1700 After use (D3) 30 0.7195 0.9663 0.0425 0.4215 4.6900

[0213] Table 12 Acne volume detection results 2

[0214]

[0215] Comedonal volume: represents the volume of comedones. The larger the value, the larger the volume of comedones.

[0216] Table 13 Acne volume detection results 3

[0217]

[0218] “ns” indicates no statistical difference, P>0.05; “*” indicates significant difference, 0.01≤P<0.05, noted as “*”; 0.001≤P<0.01, noted as “**”; p<0.001 is noted as: “***”.

[0219] After continuous use of the product for 3 days, the volume of acne decreased by 6.07% on D1 compared with the baseline value, and decreased significantly (P < 0.001) on D3 compared with the baseline value, with a decrease rate of 25.31%.

[0220] 3. Mean optical density of skin erythema

[0221] Table 14 Skin erythema optical density mean test results 1

[0222] Time quantity Mean Standard Deviation Minimum Median Maximum Before use (D0) 30 154.70 32.60 80.57 150.71 205.36 After use (D1) 30 145.37 32.24 65.39 147.00 194.00 After use (D3) 30 136.37 38.57 75.97 138.26 255.00

[0223] Table 15 Skin erythema optical density mean test results 2

[0224] Time average value Difference Rate of change Before use (D0) 154.70 - - After use (D1) 145.37 -9.33 -6.03％ After use (D3) 136.37 -18.33 -11.85％

[0225] Mean optical density of skin erythema: The smaller the value, the lighter the skin color.

[0226] Table 16 Skin erythema optical density mean test results 3

[0227]

[0228] “ns” indicates no statistical difference, P>0.05; “*” indicates significant difference, 0.01≤P<0.05, noted as “*”; 0.001≤P<0.01, noted as “**”; p<0.001, noted as: “***”.

[0229] After continuous use of the product for 3 days, the mean optical density of skin erythema decreased significantly compared with the baseline value at D1 (0.01≤P<0.05), with a decrease rate of 6.03%. The mean optical density of skin erythema decreased significantly compared with the baseline value at D3 (P<0.001), with a decrease rate of 11.85%.

[0230] 3. Self-Assessment Questionnaire

[0231] Table 17 Evaluation of the subjects using the topical gel for 3 days

[0232]

[0233]

[0234] *Evaluation indicators: 1 point is "strongly disagree", 2 points are "somewhat disagree", 3 points are "average", 4 points are "somewhat agree", 5 points are "strongly agree", Top2Box refers to the proportion of people with N≥4 points.

[0235] *Number of product statistics = 30.

[0236] Test conclusion:

[0237] After using the product for 3 consecutive days, the average redness of the subjects decreased by 4.65% compared with the baseline value on D1, and decreased significantly (P < 0.001) by 12.68% on D3.

[0238] After using the product for 3 consecutive days, the volume of acne decreased by 6.07% on D1 compared with the baseline value, and the volume of acne decreased significantly (P < 0.001) on D3 compared with the baseline value, with a decrease rate of 25.31%.

[0239] After continuous use of the product for 3 days, the mean optical density of skin erythema decreased significantly compared with the baseline value at D1 (0.01≤P<0.05), with a decrease rate of 6.03%. The mean optical density of skin erythema decreased significantly compared with the baseline value at D3 (P<0.001), with a decrease rate of 11.85%.

[0240] The results in Table 17 show that 86.67% of the subjects feel that the gel is easily absorbed, 90.00% of the subjects feel that the gel feels comfortable to use, refreshing and non-greasy, 90.00% of the subjects feel that the number of acne has been improved after using the product, 83.33% of the subjects feel that the gel can improve the redness and swelling of skin acne, 76.67% of the subjects feel that the gel has a good overall acne removal effect, 73.33% of the subjects feel that the frequency of acne is significantly reduced after using the gel, 70.00% of the subjects feel that the gel improves the skin's smoothness after use, 80.00% of the subjects feel that the gel has a redness removal effect, 73.33% of the subjects feel that the gel has a stabilizing effect on the skin, 86.67% of the subjects feel that the gel has a soothing effect after using it, 96.67% of the subjects feel that the gel product is gentle and non-irritating, 90.00% of the subjects feel that the gel product has a good overall skin feel, and 93.33% of the subjects are generally satisfied with the gel product.

[0241] In summary, using this gel product precisely for 3 days continuously is gentle and non-irritating, and has acne-removing and soothing effects.

[0242] The above are only preferred embodiments of the present invention. It should be pointed out that a person skilled in the art can make several improvements without departing from the principles of the present invention. These improvements should also be considered as within the protection scope of the present invention.

Claims

1. A Chinese medicine composition for treating acne, characterized in that: The Chinese medicine composition is prepared from the following raw materials in parts by weight: 5-30 parts of Sophora flavescens, 5-30 parts of Salvia miltiorrhiza, 5-30 parts of Chrysanthemum indici, 5-30 parts of White Peony Root, and 1-10 parts of Licorice.

2. The Chinese medicine composition according to claim 1, characterized in that: The Chinese medicine composition is prepared from the following raw materials in parts by weight: 10-25 parts of Sophora flavescens, 10-25 parts of Salvia miltiorrhiza, 10-25 parts of Chrysanthemum indici, 15-25 parts of White Peony Root, and 4-10 parts of Licorice.

3. A method for preparing the Chinese medicine composition according to claim 1 or 2, characterized in that: The following steps are involved: The raw materials are mixed with water, extracted, and the extract is concentrated to obtain fluid extract 1. Ethanol is added for alcohol precipitation, and the supernatant is taken and the ethanol is removed to obtain fluid extract 2.

4. The preparation method according to claim 3, characterized in that: The mass ratio of the total mass of the raw materials to the mass of water is 1:6-10.

5. The preparation method according to claim 3, characterized in that: The extraction temperature is 80-100° C., the extraction times are 1-2 times, and the extraction time for each time is 0.5-3 hours.

6. The preparation method according to claim 3, characterized in that: The alcohol precipitation time is 10 to 15 hours.

7. The preparation method according to claim 3, characterized in that: The mass volume ratio of the fluid extract 1 to ethanol is 1:1-3.

8. The preparation method according to any one of claims 3 to 7, characterized in that: The dosage form of the Chinese medicine composition is a gel, a patch, a solution or an ointment.

9. Use of the Chinese medicine composition according to claim 1 or 2 or the Chinese medicine composition obtained by the preparation method according to any one of claims 3 to 8 in the preparation of drugs for treating acne.

10. Use of the Chinese medicine composition according to claim 1 or 2 or the Chinese medicine composition obtained by the preparation method according to any one of claims 3 to 8 in the preparation of anti-inflammatory, oil-control or antibacterial drugs.

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