A traditional Chinese medicine composition for treating skin diseases and a preparation method and application thereof

Abstract

The application discloses a traditional Chinese medicine composition for treating skin diseases and application thereof. The traditional Chinese medicine composition is prepared from raw medicinal materials and has the following components in parts by weight: Qingdai 1 part by weight, Gypsum Fibrosum 1.6-2.4 parts by weight, Phellodendri Cortex 1.2-1.8 parts by weight, talc 2.4-3.6 parts by weight, Isatidis Folium 1.04-1.56 parts by weight, Alismatis Rhizoma 1.12-1.68 parts by weight, Poria 1.36-2.04 parts by weight, Fructus Kochiae 1.12-1.68 parts by weight, Glycyrrhizae Radix et Rhizoma 0.8-1.2 parts by weight, Radix Astragali 0.96-1.44 parts by weight. The traditional Chinese medicine composition can be used for treating skin diseases such as eczema and dermatitis, and the gel preparation has good appearance, good smearing dispersibility and stability.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of traditional Chinese medicine preparation, and particularly relates to a traditional Chinese medicine composition for treating skin diseases and a preparation method and application thereof. BACKGROUND

[0002] Eczema is a common skin disease in clinic, accounting for 15% to 30% of skin outpatient visits, and showing an upward trend. The pathogenesis of eczema is related to skin barrier dysfunction and immune system imbalance, and eczema is a high-recurrence, allergic and inflammatory skin disease, which is clinically manifested as redness, blisters, erosion and itching, and causes impaired skin barrier function and thinning of the stratum corneum.

[0003] At present, the drugs for treating eczema in clinic mainly include glucocorticoid preparations and antihistamines, and long-term medication is required. Although the symptoms can be alleviated, it is difficult to effectively cure eczema, and the symptoms are prone to relapse after drug withdrawal. Traditional Chinese medicine has a long history and rich clinical experience in the treatment of eczema. Traditional Chinese medicine has gradually become the main source for developing new eczema treatment drugs due to its good efficacy and small adverse reactions.

[0004] In the Qing Dynasty, Xu Lingtai said in Medical Source Flow Theory that the most important method of surgery is external treatment. At present, the commonly used drugs for treating eczema in clinic are glucocorticoids and antihistamines, which need long-term medication and have certain adverse reactions. Traditional Chinese medicine and natural medicine resources in China are extremely rich, and many traditional Chinese medicines have shown good therapeutic effects in the treatment of eczema, which are an important source for developing new drugs. SUMMARY

[0005] The purpose of the present application is to provide a traditional Chinese medicine composition prepared from Qingdai, Gypsum Fibrosum, Phellodendri Cortex, Talc, Isatidis Folium, Alismatis Rhizoma, Poria, Fructus Kochiae, Glycyrrhiza and Radix Rehmanniae, which can be used for treating eczema, dermatitis and other skin diseases. The traditional Chinese medicine composition can be further prepared into a gel preparation, which has good appearance, spreading dispersibility and stability.

[0006] To achieve the above purpose, the present application provides the following technical scheme:

[0007] In the first aspect, the present application provides a traditional Chinese medicine composition, which comprises the following components: Qingdai 1 part by weight, Gypsum Fibrosum 1.6-2.4 parts by weight, Phellodendri Cortex 1.2-1.8 parts by weight, Talc 2.4-3.6 parts by weight, Isatidis Folium 1.04-1.56 parts by weight, Alismatis Rhizoma 1.12-1.68 parts by weight, Poria 1.36-2.04 parts by weight, Fructus Kochiae 1.12-1.68 parts by weight, Glycyrrhiza 0.8-1.2 parts by weight, and Radix Rehmanniae 0.96-1.44 parts by weight.

[0008] Secondly, the present invention provides a traditional Chinese medicine extract, which is obtained by alcohol extraction after mixing various raw materials; the raw materials include the following components: 1 part by weight of indigo naturalis, 1.6-2.4 parts by weight of gypsum, 1.2-1.8 parts by weight of phellodendron bark, 2.4-3.6 parts by weight of talc, 1.04-1.56 parts by weight of isatis leaf, 1.12-1.68 parts by weight of alisma rhizome, 1.36-2.04 parts by weight of poria cocos, 1.12-1.68 parts by weight of kochia fruit, 0.8-1.2 parts by weight of licorice root, and 0.96-1.44 parts by weight of sanguisorba officinalis root.

[0009] According to an embodiment of the present invention, the herbal extract is obtained by alcohol extraction after mixing the raw materials; the raw materials include the following components: 1 part by weight of indigo naturalis, 2 parts by weight of gypsum, 1.5 parts by weight of phellodendron bark, 3 parts by weight of talc, 1.3 parts by weight of indigo leaf, 1.4 parts by weight of alisma rhizome, 1.7 parts by weight of poria cocos, 1.4 parts by weight of kochia fruit, 1 part by weight of licorice root, and 1.2 parts by weight of sanguisorba officinalis root.

[0010] The extraction reagent used in the alcohol extraction is 60-80% ethanol by volume, preferably 75% ethanol by volume.

[0011] The conditions for alcohol extraction are: water bath reflux, temperature 75-90℃, preferably 85℃, time 0.5-2h, preferably 1h.

[0012] In this invention, before the alcohol extraction, the mixed raw materials are first subjected to ultrafine pulverization; the ultrafine pulverization pulverizes the traditional Chinese medicine composition to 10-25 μm. After the alcohol extraction, the filtrate is concentrated by rotary evaporation; exemplarily, it is concentrated to a concentration of 1 g / ml.

[0013] Thirdly, the present invention further provides a traditional Chinese medicine preparation, wherein the traditional Chinese medicine preparation comprises the above-mentioned traditional Chinese medicine composition or traditional Chinese medicine extract.

[0014] The dosage forms of the traditional Chinese medicine preparations are gels, microemulsion gels, liposomes, creams, and ointments.

[0015] The gel comprises the following components: the above-mentioned traditional Chinese medicine composition or extract, matrix, and adjuvants;

[0016] In the gel, the mass fraction of the traditional Chinese medicine composition or the traditional Chinese medicine extract is 3-7%, preferably 5%. The mass fraction of the matrix is ​​0.8-1.2%, preferably 1%. The mass fraction of the adjuvant is 11.55-22.2%, preferably 16.78%.

[0017] The matrix is ​​one or more of carbomer, chitosan, poloxamer, and cellulose;

[0018] The additives include humectants / solvents, pH adjusters, and preservatives.

[0019] The moisturizing / solvent is glycerin or propylene glycol; the mass fraction of the moisturizing / solvent in the traditional Chinese medicine gel is 10-20%, preferably 15%.

[0020] The pH adjuster is triethanolamine or sodium hydroxide; the mass fraction of the pH adjuster in the traditional Chinese medicine gel is 1.5-2%, preferably 1.7%.

[0021] The preservative is one or more of potassium sorbate, methylparaben, and ethylparaben; the mass fraction of the preservative in the traditional Chinese medicine gel is 0.05-0.2%, preferably 0.1%.

[0022] The gel is prepared according to the following steps: the matrix and the moisturizing / solvent are added to water, allowed to stand and swell, the freeze-dried powder of the traditional Chinese medicine composition or extract, the pH adjuster and the preservative are added, and mixed to obtain the gel.

[0023] The mixing process uses a high-pressure homogenizer; the mixing conditions are: pressure 100-1200 Bar, maximum throughput 6-12 L / h, minimum throughput 30 ml.

[0024] Fourthly, the present invention further provides the application of the above-mentioned traditional Chinese medicine composition, traditional Chinese medicine extract, and traditional Chinese medicine preparation in the preparation of drugs for treating skin diseases.

[0025] In the application, the skin diseases include eczema, weeping sores, maceration sores, auricular sores, heat rash, atopic dermatitis, and contact dermatitis.

[0026] In this application, the minimum dosage of the gel is 60 mg / cm³. 2 .

[0027] Fifthly, the present invention further provides the use of the above-mentioned traditional Chinese medicine composition, traditional Chinese medicine extract, and traditional Chinese medicine preparation in the preparation of any one or more of the following drugs:

[0028] (1) Regulation of T lymphocyte CD4 + CD8 + The drug;

[0029] (2) Drugs that reduce the number of mast cells in skin tissue;

[0030] (3) Drugs that inhibit Bcl-2 protein expression;

[0031] (4) Drugs used to treat diseases related to the sphingolipid metabolism pathway, ether lipid metabolism pathway, arginine and proline lipid metabolism pathway, glycerophospholipid metabolism pathway, and glutathione metabolism pathway.

[0032] In the traditional Chinese medicine composition (HQBG) provided by this invention, Indigo Naturalis is the principal ingredient, which is salty and cold in nature, and has the functions of clearing heat and cooling blood, and relieving various heat toxins. Combined with Phellodendron Bark, it can reduce damp heat. Phellodendron Bark and Sophora Root are the assistant ingredients, which enter the heart and kidney meridians and have the functions of clearing heat and drying dampness, purging fire and detoxifying, thus helping Indigo Naturalis to exert its effects of clearing heat, detoxifying and drying dampness. Talc, Alisma Rhizome, and Poria Cocos have the functions of clearing heat and detoxifying, promoting diuresis and clearing dampness, and promoting diuresis and eliminating dampness. Gypsum has the functions of clearing heat and purging fire, promoting tissue regeneration and astringing sores. Isatis Leaf has the functions of clearing heat and detoxifying, cooling blood and eliminating spots. Kochia Fruit has the functions of clearing heat and promoting diuresis, dispelling wind and relieving itching. Sanguisorba Officinalis has the functions of cooling blood and stopping bleeding, detoxifying and astringing sores. Licorice has the function of harmonizing the various ingredients and serves as an adjuvant ingredient, which can help enhance the effects of the principal and assistant ingredients. When used together, it has the functions of clearing heat and detoxifying, drying dampness and relieving itching, promoting tissue regeneration and astringing sores, and can be used to treat eczema, dermatitis and other conditions with swelling, itching and pain and excessive exudation.

[0033] The beneficial effects achieved by this invention are as follows:

[0034] 1. The traditional Chinese medicine composition and its preparation provided by this invention can significantly improve mast cell infiltration in the skin and ear tissue of mice, reduce skin inflammation, and confirm that the related metabolic pathways may be related to glycerophospholipid metabolism; it can also effectively regulate CD4. + and CD8 + It can significantly reduce the imbalance of Bcl-2 and Bax, and has a significant effect on resisting pathogenic microorganisms. It can also significantly restore the ratio of Bcl-2 to Bax. When used externally to treat eczema, the skin lesions become thinner and eventually return to normal. This may be related to the restoration of the Bcl-2 and Bax cell regulatory system to normal, which may be one of its mechanisms of action in treating eczema. In addition, it also has the effect of clearing DPPH.

[0035] 2. The traditional Chinese medicine gel provided by this invention has good appearance, dispersibility and stability, and is non-irritating to the skin. Attached Figure Description

[0036] Figure 1 HQBG on CD4 in the dorsal skin tissue of mice with eczema + CD8 + Comparison chart showing the impact of T lymphocyte levels; in the chart: the left image represents CD4. + The image on the right represents CD8. + .

[0037] Figure 2 A comparative diagram (TB×200) showing the effects of HQBG on the histopathology of auricular skin tissue and the number of mast cells in eczema model mice. In the diagram: A: blank group; B: model group; C: Eucerin group; D: matrix group; E: HQBG-H; F: HQBG-M; G: HQBG-L; H: comparison of each group.

[0038] Figure 3A comparative diagram (TB×200) showing the effects of HQBG on the histopathology of dorsal skin tissue and the number of mast cells in eczema model mice; in the diagram: A: blank group; B: model group; C: Eucerin group; D: matrix group; E: HQBG-H; F: HQBG-M; G: HQBG-L; H: comparison of each group.

[0039] Figure 4 A comparative figure showing the effect of HQBG on Bcl-2 protein expression in the auricular skin tissue of eczema model mice (×200); In the figure: A: blank group; B: model group; C: Eucerin group; D: matrix group; E: HQBG-H; F: HQBG-M; G: HQBG-L; H: comparison of each group.

[0040] Figure 5 A comparative figure showing the effect of HQBG on Bcl-2 protein expression in the dorsal skin tissue of eczema model mice (×200); In the figure: A: blank group; B: model group; C: Eucerin group; D: matrix group; E: HQBG-H; F: HQBG-M; G: HQBG-L; H: comparison of each group.

[0041] Figure 6 A comparative figure showing the effect of HQBG on Bax protein expression in the auricular skin tissue of eczema model mice (×200); In the figure: A: blank group; B: model group; C: Eucerin group; D: matrix group; E: HQBG-H; F: HQBG-M; G: HQBG-L; H: comparison of each group.

[0042] Figure 7 A comparative figure showing the effect of HQBG on Bax protein expression in the dorsal skin tissue of eczema model mice (×200); In the figure: A: blank group; B: model group; C: Eucerin group; D: matrix group; E: HQBG-H; F: HQBG-M; G: HQBG-L; H: comparison of each group.

[0043] Figure 8 The image shows a comparison of the effects of HQBG on Bcl-2 / Bax levels in mice with eczema. In the image: Image A represents Bcl-2 / Bax in the auricle, and Image B represents Bcl-2 / Bax in the skin on the back.

[0044] Figure 9 The scatter plots of PCA under positive and negative ion modes are shown below. In the figure: A represents the positive ion mode; B represents the negative ion mode; C: blank group; M: model group; HQBG: cypress gel group; QC: quality control sample.

[0045] Figure 10The figures show the scatter plots and displacement test plots of OPLS-DA for the control group and the model group; in the figures: the upper left plot represents the scatter plot of OPLS-DA under positive ions; the upper right plot represents the displacement test plot under positive ions; the lower left plot represents the scatter plot of OPLS-DA under negative ions; the lower right plot represents the displacement test plot under negative ions; C: control group; M: model group; R 2 : Indicates the explanatory power of the constructed model for the X and Y matrices; Q 2 : Indicates the predictive power of the model.

[0046] Figure 11 The figures show the dispersion plots and displacement test plots of OPLS-DA in the model group and the HQBG group; in the figures: Figure A shows the dispersion plot of OPLS-DA under positive ions; Figure B shows the displacement test plot under positive ions; Figure C shows the dispersion plot of OPLS-DA under negative ions; Figure D shows the displacement test plot under negative ions; M: model group; HQBG: *Pterocarya stenoptera* gel group; R 2 : Indicates the explanatory power of the constructed model for the X and Y matrices; Q 2 : Indicates the predictive power of the model.

[0047] Figure 12 This diagram illustrates the effects of HQBG on metabolic pathways in a mouse model of eczema. The pathways shown are: 1. Sphingolipid metabolism; 2. Ether lipid metabolism; 3. Arginine and proline metabolism; 4. Glycerol phospholipid metabolism; 5. Glutathione metabolism; 6. Glyoxylic acid and dicarboxylic acid metabolism; 7. β-alanine metabolism.

[0048] Figure 13 This is a comparison chart showing the ability of *Platycladus orientalis* extract to scavenge DPPH free radicals. Detailed Implementation

[0049] The present invention will be further described below with reference to specific embodiments, but the present invention is not limited to the following embodiments.

[0050] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0051] Unless otherwise specified, all reagents, materials, instruments, etc. used in the following examples are commercially available.

[0052] Example 1: Preparation of Traditional Chinese Medicine Composition and Gel

[0053] 1. Instruments and Materials

[0054] 1.1 Medicines and Reagents

[0055] Indigo naturalis, talc, gypsum, Isatis tinctoria leaf, Alisma plantago-aquatica, licorice root (batch numbers 21031900, 22010900, 22031102, 19081200, 22021900, 23011000 respectively, Beijing Qiancao Traditional Chinese Medicine Pieces Co., Ltd.); Phellodendron bark, Sophora flavescens (batch numbers 221101, 220601 respectively, Beijing Bencao Fangyuan Bozhou Pharmaceutical Technology Co., Ltd.); Kochia scoparia fruit, Sanguisorba officinalis root, Poria cocos (batch numbers 890234, 390 respectively). 1224, 3907052 (Anguo City Qiao Traditional Chinese Medicine Pieces Co., Ltd.); Carbomer 980 (batch number 202208, Qingdao Tianliyuan Biotechnology Co., Ltd.); Triethanolamine (batch number 20181008, Tianjin Tianli Chemical Reagent Co., Ltd.); Glycerol, Propylene Glycol (batch numbers 20221228 and 20201237 respectively, Tianjin Fengchuan Chemical Co., Ltd.); Potassium Sorbate (batch number 20220203, Hunan Huari Pharmaceutical Co., Ltd.). Chitosan derivatives (batch number 20190018, Ruichengkang Pharmaceutical Technology Co., Ltd.), poloxamer derivatives, hydroxypropyl methylcellulose derivatives (batch numbers 20181208 and 22041503 respectively, Hunan Xinlvfang Pharmaceutical Co., Ltd.); sodium hydroxide, methylparaben, ethylparaben (batch numbers 20220009, 20190323, and 20180513 respectively, Shanghai Maclean Biochemical Technology Co., Ltd.).

[0056] 1.2 Instruments

[0057] HH-S4A electric thermostatic water bath (Beijing Kewei Yongxing Instrument Co., Ltd.), JA5003 electronic balance (Shanghai Shunyu Hengping Scientific Instrument Co., Ltd.), CHRIST freeze dryer (Shanxi Shangning Technology), RE-5299 rotary evaporator (Shanghai Yarong Biochemical Instrument Factory), HC-3018R high-speed refrigerated centrifuge (Anhui Zhongke Zhongjia Scientific Instrument Co., Ltd.); XM-PIIH stepless adjustable ultrasonic cleaner (Xiaomei Ultrasonic Instrument (Kunshan) Co., Ltd.), Scientz-150 high-pressure homogenizer (Ningbo Xinzhi Biotechnology Co., Ltd.), DX-30S small ultrafine pulverizer (Daxiang Electronic Machinery Equipment Co., Ltd.).

[0058] 2 Experimental Methods

[0059] 2.1 Preparation of Compound Extract of Phellodendron Amurense

[0060] Weigh 100g of *Phellodendron amurense* according to the prescription, grind it into ultrafine powder and mix it evenly. Add 10 times the amount of 75% ethanol and reflux in a water bath at 85°C for 1 hour. Filter it and extract it twice with 8 times the amount of 75% ethanol using the same method. Combine the two filtrates and evaporate them by rotary evaporation until they become viscous to obtain the compound extract of *Phellodendron amurense*.

[0061] 2.2 Research on the gel forming process of Cypress sphagnum moschata

[0062] 2.2.1 Selection of excipients

[0063] Commonly used matrices for gelling agents include carbomers, chitosans, poloxamers, and celluloses.

[0064] The main moisturizers are glycerin and propylene glycol.

[0065] The preservatives are mainly methylparaben, ethylparaben, potassium sorbate, etc.

[0066] pH adjusters mainly include triethanolamine and sodium hydroxide.

[0067] 2.2.2 Investigation on the effect of carbomer soaking water volume and swelling time

[0068] Take three portions of Carbomer-980, 1g each, and sprinkle them evenly into beakers containing 20, 30, and 40 times their volume of purified water, respectively. Seal the beakers and observe the swelling time of the Carbomer-980.

[0069] 2.2.3 Preparation method of *Platycladus orientalis* gel

[0070] Preparation of lyophilized powder from compound extract of *Platycladus orientalis*: The compound extract of *Platycladus orientalis* was placed in a petri dish and lyophilized in a freeze dryer. The yield of the lyophilized powder was approximately 10.76%.

[0071] Preparation of *Platycladus orientalis* gel: Weigh an appropriate amount of distilled water, add carbomer and glycerin and stir to moisten it, add an appropriate amount of distilled water, stir and let it swell naturally overnight; after complete swelling, add *Platycladus orientalis* compound extract and triethanolamine in sequence, stir evenly, then add potassium sorbate and the remaining distilled water in sequence, stir in the same direction for a long time until a clear, fine and uniform yellow transparent gel is formed. Finally, place the prepared gel in a centrifuge at 3000 r / min for 10 min to degas, and mix evenly using a high-pressure homogenizer (pressure 100-1200 Bar, maximum throughput 6-12 L / h, minimum throughput 30 ml) to obtain the gel.

[0072] Table 1 Basic formulation of gel

[0073] Formulation Dosage Sichuan Huasun compound extract 10 g of lyophilized powder was dissolved in 50 ml of water, and 30 ml of supernatant was obtained by centrifugation Carbomer 980 1.0g Glycerin 15g Potassium sorbate 0.1g Triethanolamine 1.3g Water was added to make up the weight 100g

[0074] 2.3 Establishment of Indicators and Scoring Standards

[0075] The appearance, spreadability, stability, and viscosity of the selected cypress gel were evaluated, and the detailed scoring criteria are as follows.

[0076] Table 2 Scoring Criteria

[0077]

[0078]

[0079] 2.4 Single-factor experiment

[0080] 2.4.1 Investigation of the gel matrix

[0081] Five samples of *Platycladus orientalis* gel were prepared according to the method in 2.2.3. The shapeability and viscosity of carbomer 940, carbomer 980, CMC-Na, and sodium alginate were investigated. Other preparation conditions were the same. Preliminary tests were conducted on the shapeability and viscosity of the gels to screen the types of gel matrices.

[0082] As shown in Table 3, the experimental results show that carbomer has the most ideal effect. Although CMC-Na and sodium alginate can also form gels, they are relatively weak in terms of viscosity. In particular, when the amount of CMC-Na is small, it will form a jelly-like substance.

[0083] Table 3 Results of investigation under different matrices

[0084]

[0085] 2.4.2 Screening of Carbomer Dosage

[0086] Five portions of *Platycladus orientalis* gel were prepared according to method 2.2.3, with carbomer 980 dosages of 0.8%, 1%, 1.2%, 1.4%, and 1.8%, respectively. Other preparation conditions remained the same. The experimental results are shown in Table 4. When the carbomer dosage was greater than 1.4%, the overall viscosity of the gel was high, making it difficult to coat. When the carbomer dosage was less than 0.8%, the gel's formability was poor. When the carbomer dosage was between 0.8% and 1.2%, the gel had a smooth, uniform, and delicate appearance, moderate viscosity, good formability, and was easy to coat. Therefore, a carbomer 980 dosage of 0.8% to 1.2% is preferable.

[0087] Table 4. Screening Results of Carbomer 980 Dosage

[0088] Carbomer 980 dosage (%) Appearance Viscosity Spreading 0.6 Smooth surface, uniform and delicate Slightly thin Good spreading 0.8 Smooth surface, uniform and delicate Moderate Good spreading 1 Smooth surface, uniform and delicate Moderate Good spreading 1.2 Smooth surface, uniform and delicate Moderate Good spreading 1.4 Smooth surface, uniform and delicate Moderate Good spreading 1.8 Smooth surface, uniform and delicate Thick Difficult to spread

[0089] 2.4.3 Investigation into the dosage of glycerin

[0090] Five portions of *Platycladus orientalis* gel were prepared according to method 2.2.3, with glycerol dosages of 0%, 5%, 10%, 15%, and 20%, respectively. Other preparation conditions were the same. The experimental results are shown in Table 5. The glycerol dosage of 10%-20% is appropriate.

[0091] Table 5. Screening results for glycerin dosage

[0092] Smooth surface, uniform and delicate Thick Difficult to spread Glycerin dosage (%) 0 Appearance Viscosity Spreading 5 Smooth surface, uniform and delicate Thinner Good spreading 10 Smooth surface, uniform and delicate Slightly thin Good spreading 15 Smooth surface, uniform and delicate Moderate Good spreading 20 Smooth surface, uniform and delicate Moderate Easy to spread

[0093] 2.4.4 Dosage of pH adjuster

[0094] Five portions of *Platycladus orientalis* gel were prepared according to method 2.2.3, with triethanolamine dosages of 0%, 0.5%, 1.0%, 1.5%, and 2.0%, respectively. Other preparation conditions were the same. The experimental results are shown in Table 6. The dosage of triethanolamine is preferably between 1.5% and 2%.

[0095] Table 6. Screening Results of pH Adjuster Dosage

[0096] Smooth surface, uniform and delicate Moderate Easy to spread Triethanolamine dosage (%) 0 Appearance Viscosity Spreading 0.5 Smooth surface, uniform and delicate Thin Good spreading 1 Smooth surface, uniform and delicate Slightly thin Good spreading 1.5 Smooth surface, uniform and delicate Moderate Good spreading 2 Smooth surface, uniform and delicate Moderate Good spreading 2.5 Smooth surface, uniform and delicate Moderate Good spreading

[0097] 2.4.5 Dosage of Traditional Chinese Medicine Extracts

[0098] Five portions of *Platycladus orientalis* gel were prepared according to method 2.2.3, with the amounts of herbal extract used being 5%, 8%, 10%, 12%, and 15%, respectively. Other preparation conditions remained the same. The experimental results are shown in Table 7. When the drug loading was between 3% and 7%, the gel exhibited good gelation properties, moderate viscosity, and was easy to spread. However, when the drug loading reached 9%, the gelation properties deteriorated. Therefore, a drug loading of 3% to 7% is preferable.

[0099] Table 7. Screening Results of Dosage of Traditional Chinese Medicine Extracts

[0100] Smooth surface, uniform and delicate Thick Difficult to spread Drug loading (%) 3 Appearance Viscosity Spreading 5 Yellow, smooth surface with luster, uniform and delicate Moderate Good spreading 7 Brown-yellow, smooth surface with luster, uniform and delicate Moderate Good spreading 9 Brown-yellow, smooth surface with luster, uniform and delicate Moderate Good spreading 11 Brown, smooth surface with luster, uniform and delicate Slightly thin Good spreading

[0101] 2.5 Star-point design - response surface methodology for optimizing the formulation of talc gel.

[0102] Based on the results of the single-factor experiments above, and using the comprehensive score of the gel matrix as a reference, three factors that significantly affected the gel matrix process were selected as independent variables for response surface methodology (RSM). Using Design 8.0 software, a Box-Benhnken response surface methodology was employed, employing a 3-factor, 3-level experimental design with a total of 17 experiments to investigate the combined effects of the amounts of carbomer 980 (A), glycerol (B), and triethanolamine (C) on the gel. The experimental factors and levels, results, and arrangements are shown in the table below.

[0103] Table 8 Levels of Response Surface Experiment Factors

[0104] Brown, no luster, uneven texture Thin Good spreading Factor level -1 0.8％ 10％ 1％ 0 1％ 15％ 1.5％ 1 1.2％ 20％ 2％

[0105] Table 9. Response Surface Experiment Arrangement and Results

[0106]

[0107] 2.6 Establishment of Indicator Weights

[0108] 2.6.1 Analytic Hierarchy Process (AHP)

[0109] The importance of the indicators is subjectively ranked according to viscosity = spreadability > stability > appearance, and a pairwise comparison matrix A is established, as shown in Equation 1. Column-wise normalization (each value in the matrix / the sum of its corresponding column) is performed on comparison matrix A. The sum of each row in the result is then calculated to obtain a single column result. Vector normalization is then performed on this single column result to obtain the weight coefficient matrix ω, as shown in Equation 2. Finally, the weight coefficients (W) of each indicator are obtained. j The values ​​were 0.340, 0.340, 0.227, and 0.093, respectively. To confirm the rationality of the comparison matrix, equations (3), (4), and (5) were used for consistency testing. The largest eigenvalue (λ) was found to be 4.124, the consistency index (CI) was 0.0414 (according to the table, when n=4, the consistency index RI=0.9), and the consistency ratio (CR) was 0.046<0.1, indicating that the obtained weight coefficients were effective. The relevant formulas and results are shown in equations (1) to (5).

[0110]

[0111]

[0112] CI=(λ-n) / (n-1)(4)

[0113] CR = CI / RI (5)

[0114] Note: In the formula, n is the number of indicators, i is the i-th indicator, i = 1, 2, 3...

[0115] 2.6.2 CRITIC Method

[0116] The index values ​​in Table 5 were standardized using the formula [(index value - minimum value) / (maximum value - minimum value)] × 100%. The correlation coefficient matrix C was obtained using SPSS 25.0 (the indexes in the matrix, from left to right and top to bottom, are viscosity, spreadability, release rate, and drug loading). The contrast intensity (σ) was then calculated. j ), conflict (R) j ), comprehensive weight (C) j ) and weight (W) j The final weight coefficients for each indicator are 0.267, 0.214, 0.323, and 0.196, respectively. The relevant results and formulas are shown in equations (6) to (10) and Table 10.

[0117]

[0118]

[0119]

[0120] H j =σj R j (9)

[0121]

[0122]

[0123] Note: B ij Represents the sample values ​​corresponding to factors i and j in matrix B; B i Let represent the mean of the sample corresponding to factor i in matrix B; i, j = 1, 2, 3, ...

[0124] Table 10. Relevant parameters for calculating the weights of molding process parameters using the CRITIC method.

[0125] Carbomer 980 (A / %) Glycerin (B / %) Triethanolamine (C / %) Parameter Viscosity [["b j ]]> 0.236 0.230 0.368 0.226 [R j ]]> 3.116 2.563 2.420 2.392 H j ]]> 0.734 0.588 0.890 0.540 [WC j ]]> 0.267 0.214 0.323 0.196

[0126] 2.6.3 AHP-CRITIC Hybrid Weighted Method

[0127] The repetition weight coefficient (W) of the above indicators is obtained by equation (11). f The corresponding values ​​were 0.356, 0.285, 0.288, and 0.072. Correlation analysis using SPSS 20.0 software showed that the correlation coefficient between AHP and CRITIC was 0.463, which was not statistically significant (P > 0.05). This indicates that the information reflected is not additive. Considering both subjective and objective factors, the AHP-CRITIC weighted method is more scientific, reasonable, and stable than the single weighting method. Therefore, this study ultimately used this method to determine the indicator weights and calculate the comprehensive score.

[0128] 2.6.4 Comparison of Comprehensive Evaluation Results

[0129] The experimental data were weighted using the calculation results of three methods: AHP, CRITIC, and AHP-CRITIC. The results are shown in the table below. Correlation analysis using SPSS 20.0 software showed that the correlation coefficient between AHP and CRITIC was 0.463, which was not statistically significant (P > 0.05). This indicates that the information reflected is not additive. Considering both subjective and objective factors, the AHP-CRITIC weighted method is more scientific, reasonable, and stable than a single weighting method. Therefore, this study ultimately used this method to determine the indicator weights and calculate the comprehensive score.

[0130] Table 11. Comprehensive scoring results of three weighting methods for *Platycladus orientalis* gel.

[0131]

[0132]

[0133] 2.7 Model Fitting and Validation Experiments

[0134] 2.7.1 Model Fitting

[0135] Based on the experimental results, the regression equation was fitted using Design-Expert 12, resulting in a binomial equation:

[0136] Y=81.331+1.172A-0.026B-0.945C-0.972AB+2.632AC+0.424BC-2.449A 2 -1.824B 2 -1.469C 2

[0137] (R 2 =0.9634, P<0.0001), from R 2 As can be seen from the P-value, the regression equation has a good fit and small experimental error, and can be used for prescription prediction of cypress gel matrix. The results of the analysis of variance are shown in Table 12.

[0138] Table 12 Results of Analysis of Variance

[0139] Spreading Stability Appearance Source F P Sum of squares 96.76369 9 10.75152 5.414492 0.0002 df 10.99571 1 10.99571 5.537463 0.0508 Mean square 0.005513 1 0.005513 0.002776 0.9595 Model 7.14609 1 7.14609 3.598788 0.0996 A-carbomer 1.422056 1 1.422056 0.716151 0.4254 B-glycerin 31.29284 1 31.29284 15.75915 0.0054 C-triethanolamine 0.936056 1 0.936056 0.4714 0.5144 A 2 ]]> 15.87043 1 15.87043 7.992388 0.0255 B 2 ]]> 10.15106 1 10.15106 5.112099 0.0582 C 2 ]]> 14.26213 1 14.26213 7.182443 0.0315

[0140] 2.7.2 Response Surface Optimization and Prediction

[0141] Based on the two-dimensional contour map and the three-dimensional effect surface map, the optimal prescription ratio is set when Y is maximized: carbomer 1.06, glycerol 15.20, triethanolamine 1.48, at which point the overall score is 81.51.

[0142] 2.7.3 Optimal Process Verification Experiment

[0143] Considering the actual situation, the optimal process was ultimately determined to be 1.0g of carbomer, 15g of glycerol, and 1.5g of triethanolamine. Three validation tests were conducted according to the above matrix formulation, and the deviation value was calculated [deviation value = (predicted value - measured value) / predicted value × 100%]. The results showed that the deviations between the predicted and measured values ​​were 1.63%, 1.30%, and 1.23%, respectively, with an average relative error of 1.39%. This indicates that the established equation has good predictive ability and can accurately predict the optimal ratio of the *Platycladus orientalis* gel matrix.

[0144] Example 2: Study on the effect of Huaqingbai gel in treating eczema

[0145] The effects of Huaqingbaigel (HQBG) on the improvement of eczema model mice induced by 2,4-dinitrochlorobenzene (DNCB) acetone olive oil solution and its regulatory effects on differentially expressed metabolites in serum were investigated using liquid chromatography-mass spectrometry (LC-MS) metabolomics. The potential mechanism of HQBG in the treatment of eczema was analyzed, laying a foundation for the clinical application of HQBG.

[0146] 1. Materials

[0147] 1.1 Laboratory Animals

[0148] Fifty-six SPF-grade KM mice (half male, half female), weighing (40g ± 4g), were purchased from Spiford (Beijing) Biotechnology Co., Ltd. License No.: SCXK(Beijing)2019-0032. Animal Experiment Ethics Approval No.: 2022DW328. All mice were kept in an environment with controlled temperature (20-26℃) and humidity (40-70%), with free access to food and water, and a 12h / 12h light / dark cycle.

[0149] 1.2 Drugs and Reagents

[0150] 2,4-Dinitrochlorobenzene (DNCB, batch number LD51W29, Beijing Bailingwei Technology Co., Ltd.); Eucerin (batch number 21111004, Tianjin Jinyao Pharmaceutical Co., Ltd.). CD4 + CD8 + Enzyme-linked immunosorbent assay (ELISA) kits (batch numbers 20220716 and 20220722, respectively, Vanco Technology Shanghai Co., Ltd.). Toluidine blue staining solution (batch number 20230216, Beijing Solarbio Technology Co., Ltd.). Xylene, neutral resin (batch numbers 10023514 and 10004231, respectively, Sinopharm Chemical Reagent Co., Ltd.), general tissue fixative and blocking solutions (batch numbers G1101 and G1208-5ML, respectively, Wuhan Saiweier Biotechnology Co., Ltd.); Bax antibody and Bcl-2 antibody (batch numbers BB11156343 and BB03235878, respectively, Beijing Bio-Sens Biotechnology Co., Ltd.); hematoxylin staining solution (batch number PH0516, Fuzhou Feijing Biotechnology Co., Ltd.); HRP chromogenic solution (DAB, batch number 20230825, Kaiji Biotechnology Development Co., Ltd.); methanol (batch number 20220315, Thermo Fisher Scientific China Co., Ltd.).

[0151] 1.3 Main Instruments

[0152] UHPLC-Q Exactive high-resolution mass spectrometer, Ulti-Mate 3000 ultra-high performance liquid chromatograph (Thermo Fisher Scientific (China) Co., Ltd.), Acquity UPLC HSS T3 (2.1×100mm, 1.8μm) column (Waters Corporation, USA), Multiskan FC microplate reader, AC8 plate washer (Thermo Fisher Scientific Shanghai Instrument Co., Ltd.), WD-1000 nitrogen evaporator (Agilent Technologies Inc.), HC3018R high-speed refrigerated centrifuge (Anhui Zhongke Zhongjia Scientific Instrument Co., Ltd.), Axioscope 5 microscope (Carl Zeiss Shanghai Management Co., Ltd.).

[0153] 2 Experimental Methods

[0154] 2.1 Model preparation, grouping, and administration

[0155] Fifty-six KM mice were randomly divided into seven groups: a control group (Con.), a model group (Mod.), an evodia group (Hyd., 1.5 mg / cm³), and a control group (Hyd., 1.5 mg / cm³). 2 ), matrix group (Mat.), and high-dose group of *Pterocarya stenoptera* gel (HQBG-H, 240 mg / cm³). 2 ), medium-dose group of cypress gel (HQBG-M, 120mg / cm) 2 Low-dose group of cypress gel (HQBG-L, 60 mg / cm³) 2 ).

[0156] After acclimatizing the mice for one week, hair was removed from the backs of the mice using depilatory cream before the experiment, covering an area of ​​approximately 2×2cm. 2 On day 1 of the experiment, the control group received a 100 μL acetone-olive oil solution (acetone:olive oil = 4:1) applied topically to the back. All other groups received a 7% DNCB solution (100 μL) applied to the hairless area to induce sensitization. On day 6, dermatitis was induced by applying a 0.7% DNCB-acetone-olive oil solution (20 μL) to the back and behind the ears of the mice, with sensitization occurring every two days. Except for the normal and model groups, which received distilled water, the other groups received the corresponding drug based on the area treated, for 12 consecutive days. Topical application of the drug began on the day of stimulation. If DNCB stimulation was required on the day of administration, the interval between administration and stimulation was 2 hours. Successful modeling was indicated by the appearance of dry, itchy, red, swollen, or thickened skin in the mice.

[0157] 2.2 T lymphocytes CD4 + CD8 + Horizontal detection and analysis

[0158] Skin lesions from the backs of mice in each group were collected at -80℃, thawed, and 0.1g was weighed into a 2ml centrifuge tube. 0.9ml of physiological saline was added, and the tissue was homogenized. After complete tissue disruption, the tissue was vortexed for 1 minute, centrifuged, and the supernatant was collected. CD4+ in T lymphocytes of the skin tissue was detected using ELISA. + and CD8 + The levels were determined according to the instructions of the relevant reagent kits.

[0159] 2.3 Histopathological Experiments

[0160] Skin lesions from mice in each group were collected, fixed with paraformaldehyde, dehydrated with ethanol, cleared with xylene, embedded and fixed in paraffin, sectioned, stained with toluidine blue (TB), mounted with neutral resin, and the pathological changes of the dorsal skin and auricular tissue were observed under a microscope. Six fields of view were randomly selected from each group, and the number of mast cells in each field of view was measured using Image-Pro Plus 6.0 software, followed by statistical analysis.

[0161] 2.4 Immunohistochemical detection

[0162] Paraffin sections of mouse skin were melted in a 60°C oven, followed by xylene dewaxing and graded ethanol dehydration. Tissue sections were placed in an antigen retrieval buffer (pH 6.0) and microwaved for 5 min for antigen retrieval. They were then incubated with primary antibodies (Bcl-2, 1:150 dilution) and Bax (1:150 dilution) and their corresponding secondary antibodies, respectively. DAB staining and hematoxylin counterstaining were performed, followed by mounting and observation.

[0163] Under a microscope, six identical planar slices were randomly selected from each specimen, and the average optical density (OD) value of the positive substance expression area was calculated using image-proplus6.0 image analysis software.

[0164] 2.5 LC-MS analysis of serum metabolomics in each group of mice

[0165] 2.5.1 Liquid chromatography-mass spectrometry (LC-MS) sample collection and processing

[0166] Blood was collected from mice, and after resting at room temperature for 0.5 h, the mice were irradiated at 1500 r / min. -1Centrifuge for 20 min, collect serum and store at -80℃ for later use. Thaw at 4℃ before use, take 100 μL of serum, add 3 times pre-cooled chromatographic grade methanol and repeatedly pipette, centrifuge at 1300 r / min for 5 min at 4℃, dry under nitrogen, reconstitute with 100 μL of mass spectrometry grade methanol before loading, centrifuge, and collect the supernatant. Accurately pipette 10 μL of each sample into a centrifuge tube, vortex to mix, and prepare quality control (QC) samples. Filter each sample through a 0.22 μm organic filter membrane into an autosampler vial for UPLC-Q-TOF-MS analysis.

[0167] 2.5.2 Chromatographic-Mass Spectrometry Conditions

[0168] Chromatographic column: Acquity UPLC HSS T3 (2.1×100mm, 1.8μm), injection volume: 5μL, flow rate: 0.3mL / min, column temperature: 40℃. Mobile phase: acetonitrile (A), 0.1% formic acid-water (B); gradient elution method: 0-0.5min (A) 5%, 0.5-1.5min (A) 5%-15%, 1.5-4.5min (A) 15%-30%, 4.5-6min (A) 30%-60%, 6-9min (A) 60%-70%, 9-12min (A) 70%-100%, 12-13min (A) 100%, 13-13.5min (A) 100%-5%, 13.5-16min (A) 5%. MS recording of total ion current (TIC) chromatograms, electrospray ionization (ESI) source, simultaneous scanning mode for positive and negative ions. Spray voltage 3.2 kV, sheath gas flow rate 40 arb, auxiliary gas flow rate 5 arb, auxiliary gas heating temperature 350 °C; ion transmission tube temperature 320 °C, S-Lens RF Level 50 V, scan range 100–1000 m / z, collision energy 30 eV.

[0169] 2.5.3 Data Processing

[0170] Metabolomics data were analyzed using Compound Discovery software, obtaining detailed information including retention time, m / z value, and peak area for each ion peak. Peak areas were normalized. Principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) were performed using SIMCA 14.1 software, and the model was permuted using 200 permutations. Differential metabolites were screened based on VIP>1 and P<0.05 in the S-plot. Potential metabolites were identified using the Human Differential Metabolome Database (HMDB). Cluster analysis and pathway enrichment analysis of differential metabolites were performed using the MetaboAnalyst 5.0 online analysis platform.

[0171] 2.6 Statistical Analysis

[0172] Data analysis and image processing were performed using Graphpad Priam 9.0 and Image-Pro Plus 6.0 software. Quantitative data were used... The data comparison among multiple groups was performed using one-way ANOVA, and P < 0.05 indicated that the differences were statistically significant.

[0173] 3 Results

[0174] 3.1 T lymphocytes CD4 + CD8 + Horizontal analysis results

[0175] ELISA assay of HQBG on CD4 levels in the dorsal skin of eczema model mice + CD8 + The effect of T lymphocyte levels, results are shown in AB Compared with the control group, the model group mice had higher levels of CD4+ in their skin tissue. + CD8 + The level of T lymphocytes was significantly increased, with a highly significant difference (P < 0.01); the Hyd group and each HQBG administration group significantly reduced the CD4+ level in T lymphocytes in the skin tissue of eczema model mice. + CD8 + The levels were statistically significant (P < 0.01). In conclusion, HQBG can significantly downregulate CD4. + CD8 + At that level, it has a significant effect against pathogenic microorganisms.

[0176] 3.2 Histopathological Experimental Results

[0177] Mast cells in each group of skin lesions were stained using TB staining. The results of TB staining of the auricle are as follows: AC The control group showed intact tissue structure, clear layers, and no mast cell infiltration. In contrast, the model group mice exhibited extensive mast cell infiltration in their ear tissues after repeated stimulation with DNCB. Intervention with various doses of Hyd and HQBG reduced mast cell infiltration. BC (H) indicates that, compared with the blank group, the number of mast cells in the auricular tissue of the model group was significantly increased (P<0.01); compared with the model group, the Hyd group and each dose group of HQBG significantly reduced the number of mast cells in the skin (P<0.01), thereby alleviating the skin lesions of the auricular tissue of mice induced by DNCB.

[0178] HE staining results of the back skin are as follows Figure 1In the control group, the epidermal cells on the back of mice were arranged in an orderly manner, the tissue thickness was normal, and mast cell infiltration was obvious. Compared with the control group, the model group induced by DNCB stimulation showed severe hyperkeratosis of the back skin, with obvious mast cell infiltration. Compared with the model group, mast cell infiltration in the skin tissue of the Hyd group and each HQBG administration group was significantly reduced after intervention. Figure 2 (H) indicates that, compared with the control group, the number of mast cells in the back skin tissue of the model group was significantly increased (P < 0.01); compared with the model group, the Hyd group and each dose group of HQBG significantly reduced the number of mast cells in the back skin tissue (P < 0.01). Therefore, HQBG can significantly improve the pathological changes in the DNCB-induced eczema model.

[0179] 3.3 Immunohistochemical detection results

[0180] 3.3.1 Results of Bcl-2 protein expression

[0181] After staining, Bcl-2 protein expression in the auricle was found in the cytoplasm of keratinocytes and lymphocytes, appearing as brownish-yellow granules. The staining results are shown in the attached image. Figure 2 In the control group, Bcl-2 protein expression was weakly positive, and the nuclei and cell membranes of keratinocytes and lymphocytes were intact without damage, with brownish-yellow granules visible in the cells. After repeated stimulation with DNCB, Bcl-2 protein expression in the model group was strongly positive, with a large number of keratinocytes and lymphocytes visible, some fragmented, and many brownish-yellow granules visible. After intervention in the Hyd group and various doses of HQBG, Bcl-2 protein expression was weakly positive, the number of keratinocytes and lymphocytes decreased, and the amount of brownish-yellow granules in the cytoplasm decreased.

[0182] Depend on Figure 3 (H) indicates that, compared with the blank group, the OD value in the auricular tissue of the model group was significantly increased (P<0.01); compared with the model group, the OD values ​​of the Hyd group and each dose group of HQBG were significantly reduced (P<0.05).

[0183] The results of staining the skin on the back are shown in the figure. Figure 3 In the control group, Bcl-2 protein expression was weakly positive, the cell nucleus and cell membrane were intact and undamaged, and brownish-yellow granules were visible in the cells. After repeated stimulation with DNCB, Bcl-2 protein expression in the model group was strongly positive, with a large number of brownish-yellow granules visible and some cells fragmented. After intervention in the Hyd group and various doses of HQBG, Bcl-2 protein expression became positive, and the brownish-yellow granules in the cytoplasm were significantly reduced.

[0184] Depend on Figure 4(H) indicates that, compared with the blank group, the OD value in the back skin tissue of the model group was significantly increased (P<0.05); compared with the model group, the OD values ​​of the Hyd group and each dose group of HQBG were significantly reduced (P<0.05).

[0185] 3.3.2 Bax protein expression results

[0186] After staining, Bax protein expression was found in the cytoplasm of keratinocytes and lymphocytes, appearing as brownish-yellow granules. The staining results are shown in the figure. Figure 4 In the control group, Bax protein expression was positive, and the nuclei and cell membranes of keratinocytes and lymphocytes were intact without damage, with visible brownish-yellow granules. After repeated stimulation with DNCB, Bax protein expression in the model group was weakly positive, with a large number of keratinocytes and lymphocytes, some of which were fragmented, and the visible brownish-yellow granules decreased. After intervention in the Hyd group and each dose of HQBG, Bcl-2 protein expression was positive, the nuclear membrane was intact, and the amount of brownish-yellow granules in the cytoplasm increased.

[0187] Depend on Figure 5 (H) indicates that, compared with the blank group, the OD value in the auricular tissue of the model group was significantly reduced (P<0.01); compared with the model group, the OD values ​​of the Hyd group and each dose group of HQBG were significantly increased (P<0.05).

[0188] The results of staining the skin on the back are shown in the figure. Figure 5 In the control group, Bax protein expression was positive, the nuclear membrane was intact and undamaged, and brownish-yellow granules were visible. After repeated stimulation with DNCB, Bax protein expression in the model group was weakly positive, the brownish-yellow granules decreased, and some cells fragmented. After intervention in the Hyd group and various doses of HQBG, Bax protein expression was positive, and the brownish-yellow granules in the cytoplasm increased significantly.

[0189] Depend on Figure 6 (H) indicates that, compared with the blank group, the OD value in the back skin tissue of the model group was significantly reduced (P<0.05); compared with the model group, the OD value of each dose group of Hyd group and HQBG group was significantly increased (P<0.05).

[0190] 3.3.3 Bcl-2 / Bax ratio

[0191] The results of the Bcl-2 / Bax ratio in the skin tissue of the auricle and back are shown in the following figures. Figure 6 (A) and (B) Compared with the blank group, the Bcl-2 / Bax value in the tissue of the model group was significantly increased (P<0.01); compared with the model group, the Bcl-2 / Bax value in each dose group of Hyd group and HQBG was significantly reduced (P<0.01).

[0192] 3.4 LC-MS results of metabolomics analysis of serum from each group of mice

[0193] 3.4.1 PCA Analysis

[0194] Multivariate statistical analysis was used to further identify differentially expressed metabolites among the groups. PCA analysis revealed the spatial distribution of serum samples from each group. Results are shown below. Figure 7 In the positive ion model, R 2 X = 0.306, Q 2 =0.127, R in the negative ion model 2 X = 0.344, Q 2 =0.158, indicating a relatively clear distinction between the model group and the normal group. The sclerotium affine gel group was closer to the normal group, which may suggest that the metabolic disorders in the eczema model mice were reversed after drug intervention.

[0195] 3.4.2 OPLS-DA Analysis

[0196] Establish supervised OPLS-DA analysis, such as Figure 7 As shown. The samples in the blank group and the model group are clearly distinguishable; in the positive ion model, R... 2 X = 0.314, R 2 Y = 1, Q 2 =0.697, R in the negative ion model 2 X = 0.343, R 2 Y = 1, Q 2 =0.778, indicating that the model has good fit and predictive ability. A permutation test was performed on the model with n=200. Figure 8 (B) and Figure 9 (D) The model's slope was found to be relatively large and intersected the ordinate on the negative half-axis, indicating that the model was not overfitting and was reliable. Based on the VIP>1 criterion and the independent samples t-test (P<0.05), differentially differentiated metabolites were screened, ultimately yielding 862 differentially differentiated metabolites with significant differences.

[0197] The OPLS-DA results for the model group and the HQBG group are as follows: Figure 10 As shown, effective separation can be achieved between the two groups of samples, and R in the positive ion model 2 X = 0.338, R 2 Y = 1, Q 2 =0.849, R in the negative ion model 2 X = 0.33, R 2 Y = 1, Q 2 =0.859, R 2 Y and Q 2All values ​​are greater than 0.5, indicating that the model is well-established and suitable for the analysis and screening of differential metabolites. A permutation test was performed on 200 random permutations of the model, revealing a large slope that intersects the negative half-axis of the ordinate, indicating that the model is not overfitted and is reliable. Figure 10 (B) and Figure 10 (D). Based on the VIP>1 criterion, an independent samples t-test was conducted (P<0.05), ultimately identifying 658 differentially expressed metabolites.

[0198] 3.4.3 Screening and Identification of Differential Metabolites

[0199] The control group and model group yielded 862 differentially expressed metabolites, while the model group and HQBG group yielded 658 differentially expressed metabolites, with a total of 233 metabolites in the intersection. A total of 25 differentially expressed metabolites were identified in the HMDB database.

[0200] Metabolite heatmap analysis using MetaboAnalyst 5.0 showed that the model group roughly clustered into one group, while the normal group and the HQBG group roughly clustered into another. Compared with the model group, HQBG administration significantly regulated the metabolism of eczema model mice, suggesting that these differentially metabolites may be closely related to the pathway of HQBG treatment for eczema.

[0201] Table 13 Serum Differential Metabolites

[0202]

[0203]

[0204] 3.4.4 Differential Metabolite Pathway Enrichment Analysis

[0205] Twenty-five differentially expressed metabolites were input into the MetaboAnalyst 5.0 database to construct metabolic pathways. Seven pathways related to HQBG-induced eczema in mice were identified, and five pathways with impact>0 were selected as potential target pathways. (See attached data). Figure 11 These are sphingolipid metabolism, ether lipid metabolism, arginine and proline lipid metabolism, glycerophospholipid metabolism, and glutathione metabolism, respectively.

[0206] Example 3: Determination of the antioxidant capacity of *Platycladus orientalis* extract

[0207] 1. Materials and Instruments

[0208] 1.1 Experimental Reagents

[0209] Anhydrous ethanol; 1-Diphenyl-2-trinitrophenylhydrazine (DPPH, batch number CD29142801, Beijing Cooler Technology Co., Ltd.); 2-Diazon-bis(3-ethyl-benzothiazole-6-sulfonic acid)-diammonium salt (ABTS, batch number 30931670, Shanghai Yuanye Biotechnology Co., Ltd.); Potassium persulfate (batch number 20160905, Tianjin Fengchuan Chemical Reagent Technology Co., Ltd.); Ferrous sulfate (batch number 2016222, Tianjin Beichen Fangzheng Reagent Factory); Hydrogen peroxide (batch number 20180321, Tianjin Beilian Fine Chemicals Development Co., Ltd.); Salicylic acid (batch number C12173991, Shanghai Maclean Biochemical Technology Co., Ltd.).

[0210] 1.2 Instruments

[0211] HH-6 digital display constant temperature water bath (Changzhou Ronghua Instrument Manufacturing Co., Ltd.); EX125ZH type 0.0001 g electronic balance (Ohaus Instruments Changzhou Co., Ltd.); Ultra-3600 ultraviolet-visible spectrophotometer (Beijing Puyuan Precision Technology Co., Ltd.).

[0212] 2. Test Methods

[0213] 2.1 Preparation of *Platycladus orientalis* extract

[0214] Same as the extract of *Platycladus orientalis* in Example 1.

[0215] 2.2 Determination of the antioxidant activity of Cypress sapwood

[0216] 2.2.1 Determination of the ability of *Platycladus orientalis* extract to remove DPPH free radicals

[0217] Weigh 3.95 mg of DPPH and dissolve it in a 100 mL volumetric flask to prepare a 0.1 mmol / L DPPH solution. Store the solution away from light.

[0218] Mix 1.5 mL of 0.1 mmol / L DPPH solution with 1.5 mL of extracts of different concentrations of *Platycladus orientalis*, and let stand in the dark for 30 min. Use anhydrous ethanol as a blank control and measure the absorbance A1 at 517 nm. Perform three parallel measurements.

[0219] Blank control: 1.5 mL of anhydrous ethanol and 1.5 mL of different concentrations of *Platycladus orientalis* extract were mixed evenly and placed in the dark for 30 min. The absorbance of the mixture was measured at 517 nm using anhydrous ethanol as a blank control. The absorbance was measured in parallel three times.

[0220] Negative control: Mix 1.5 mL of DPPH solution with 1.5 mL of anhydrous ethanol, incubate in the dark for 30 min, and then measure the absorbance A0 at 517 nm using anhydrous ethanol as a blank control. Use the corresponding VC as a control. Calculate the DPPH free radical scavenging ability of the *Platycladus orientalis* extract according to the following formula:

[0221] Clearance rate (%) = [1 - (A1 - A2) / A0] × 100%

[0222] 2.2.2 Determination of the ability of *Platycladus orientalis* extract to remove ABTS free radicals

[0223] Mix 0.2 mL of 7.4 mmol / L ABTS solution with 0.2 mL of 2.6 mmol / L potassium persulfate solution and let stand at room temperature in the dark for 12 h to obtain the ABTS reaction solution. Then dilute with anhydrous ethanol approximately 40 times to obtain an absorbance of 0.7 ± 0.02 at 734 nm. Take 2.4 mL of the ABTS reaction solution and shake with 0.6 mL of 95% ethanol for 10 s, let stand in the dark for 6 min, and measure the absorbance at 734 nm (A0). Take 2.4 mL of the ABTS reaction solution and shake with 0.6 mL of the sample solution for 10 s, let stand in the dark for 6 min, and measure the absorbance at 734 nm (A). Use the corresponding VC as a control. Calculate the scavenging ability of *Platycladus orientalis* extract against ABTS free radicals according to the following formula:

[0224] Clearance rate (%) = (A0 - A) / A0 × 100%

[0225] 2.2.3 Determination of the ability of *Platycladus orientalis* extract to scavenge hydroxyl radicals

[0226] 0.6 mL of 7.5 mmol / L ferrous sulfate, 0.6 mL of 7.5 mmol / L hydrogen peroxide, 0.6 mL of 7.5 mmol / L salicylic acid, and 2 mL of different concentrations of *Platycladus orientalis* extract were incubated in a water bath at 37°C for 30 min. The absorbance (A1) was measured at 510 nm. When 0.6 mL of distilled water was used instead of hydrogen peroxide, the OD value was measured as A2. When 2 mL of distilled water was used instead of different concentrations of *Platycladus orientalis* extract, the absorbance value was measured as A0. Using the corresponding vitamin C as a control, the scavenging ability of *Platycladus orientalis* extract against hydroxyl radicals was calculated using the following formula:

[0227] Clearance rate (%) = [1 - (A1 - A2) / A0] × 100%

[0228] 3 Results

[0229] Determination of the ability of *Platycladus orientalis* extract to remove DPPH free radicals: DPPH is a stable nitrogen-centered free radical. If the test substance can remove it, it indicates that it has the effect of scavenging free radicals. DPPH appears purple in ethanol solution and has maximum absorption at 517 nm. In the presence of free radical scavengers, the lone pair electrons of DPPH are paired, making its color lighter and the absorbance at the maximum absorption wavelength decreases. In the experiment, the scavenging rate is used to represent the ability of the test substance to remove free radicals. The higher the scavenging rate, the stronger its antioxidant capacity.

[0230] Depend on Figure 11 It was found that both *Platycladus orientalis* extract and vitamin C exhibit DPPH scavenging activity, with the scavenging rate increasing with increasing concentration. However, the scavenging ability significantly increased from 1 mg / ml to 4 mg / ml. From 4 mg / ml to 10 mg / ml, the increase in scavenging ability slowed down. Vitamin C's ability to scavenge DPPH increased slightly with increasing concentration, but the change was not significant. Overall, *Platycladus orientalis* extract has a significant DPPH scavenging effect.

[0231] Example 4: Experimental protocol for the skin irritation of cypress gel on rabbits.

[0232] 1. Materials

[0233] 1.1 Drugs and Reagents

[0234] HQBG gel and blank matrix.

[0235] 1.2 Laboratory Animals

[0236] Healthy young domestic rabbits, half male and half female, weighing 0.8–1.2 kg; healthy young guinea pigs, half male and half female, weighing 150–200 g. Animals were housed individually with free access to food. The experiment began after one week of acclimatization.

[0237] 2 Experimental Methods

[0238] 2.1 Local skin irritation in young rabbits

[0239] 2.1.1 Grouping

[0240] The single-dose intact skin test group (including the HQBG test drug group and the blank matrix negative control group, a total of 2 groups, 6 young rabbits in each group) and the broken skin test group (including the HQBG test drug group and the blank matrix negative control group, a total of 2 groups, 6 young rabbits in each group).

[0241] The test groups included two groups: the intact skin test group (comprising the HQBG test drug group and the blank matrix negative control group, with 6 rabbits in each group) and the damaged skin test group (comprising the HQBG test drug group and the blank matrix negative control group, with 6 young rabbits in each group).

[0242] 2.1.2 Administration methods for intact and broken skin

[0243] To investigate the irritant response of intact rabbit skin, six healthy rabbits (half male and half female) were used. Two symmetrical patches of hair were removed from each side of their backs, each approximately 3cm x 3cm in size. The experiment employed a self-controlled left-right side method, where 0.5g / kg of HQBG was applied to the skin on the left back, and 0.5g / kg of the matrix was applied to the skin on the right back.

[0244] To investigate the irritation response of rabbits to broken skin, six healthy rabbits (half male and half female) were used. Two symmetrical patches of hair were removed from each side of their backs, each approximately 3cm x 3cm in size. Using a sterile No. 8 needle, 2cm diameter "#"-shaped abrasions were made, ensuring only the epidermis was pierced and the dermis was not damaged, aiming for slight bleeding. The experiment used a self-controlled left-right comparison method: 0.5g / kg of HQBG was applied to the skin on the left back, and 0.5g / kg of the base was applied to the skin on the right back.

[0245] 2.1.3 Single Dosage Method

[0246] For the single-dose skin irritation test, one group of intact hairless skin and one group of broken skin (6 animals per group) were used. 0.5 g / kg of the test substance was evenly applied to the left side of the hairless area, then covered with one layer of plastic wrap and two layers of gauze. The entire back and abdomen were then secured with a latex glove with the finger cots cut off. A blank matrix was applied to the right side as a negative control. After application, the skin was fixed with two layers of gauze and one layer of non-irritating adhesive tape for 6 hours. After 6 hours, the test drug was removed, and the application site was washed with 0.9% sodium chloride solution. Before administration and at 0.5 h, 1 h, 24 h, 48 h, and 72 h, the animals were observed for erythema and edema, and for pigmentation, bleeding points, rough skin, or thinning of the skin at the application site. The appearance and resolution times were recorded.

[0247] 2.1.4 Multiple Dosing Method

[0248] For the repeated-dose skin irritation test, one group (6 animals per group) of intact hairless skin and one group of broken skin were used. The administration method was the same as for single-dose administration, but the same medication was applied to the same site for 14 consecutive days at the same time each time. After each application, the area was fixed with two layers of gauze and one layer of non-irritating adhesive tape for 6 hours. After fixation, the test drug was removed and the application site was cleaned with warm water. The medication was applied continuously for 14 days. Before each administration and at 0.5h, 1h, 24h, 48h, and 72h after the last administration, the animals were observed for erythema and edema, as well as for pigmentation, bleeding points, rough skin, or thinning of the skin at the application site. The time of appearance and disappearance was recorded.

[0249] 2.1.5 Stimulus Intensity Evaluation

[0250] The skin at the experimental site was observed for erythema and swelling before administration and at 0.5h, 1h, 24h, 48h, and 72h after administration. Evaluation criteria:

[0251] Table 14 Skin Irritation Response Scoring Criteria

[0252]

[0253] Table 15 Mild Skin Irritation Grading

[0254] Figure 11 Figure 12 0.00～0.49 Figure 13 0.50～1.99 Integral mean 2.00～5.99 Degree 6.00～8.00 No irritation Mild irritation Moderate irritation Strong irritation

[0255] The average score for the stimulus response = (total score for erythema formation + total score for edema formation) / number of animals.

[0256] 2.1.6 Observation Indicators

[0257] Before daily administration and at 0.5h, 1h, 24h, 48h, and 72h after the last administration, observe the animals for erythema and edema reactions, and note any pigmentation, petechiae, rough skin, or thinning of the skin at the application site. Record the time of appearance and resolution. Erythema and edema are scored, and the average daily stimulation score for each rabbit is calculated during the observation period.

[0258] 3 Experimental Results

[0259] In both the HQBG test drug group and the blank matrix-negative control group, rabbits in the intact skin group, regardless of whether they received a single or multiple doses, showed mild erythema and edema on the treated side during the administration and recovery observation periods. This may be related to individual differences, with skin irritation scores ranging from 0 to 0.33, indicating no skin irritation. No erythema, edema, or other irritation symptoms were observed on the matrix-negative control side; the skin irritation score was 0, indicating no skin irritation. The irritation response scores for single application to intact skin in rabbits are shown in Table 16, and the irritation response scores for multiple application to intact skin in rabbits are shown in Table 17.

[0260] In both the HQBG test drug group and the blank matrix negative control group, regardless of whether the drug was administered once or multiple times, a few rabbits in the damaged skin group developed transient, mild erythema on the administered side and the control side after drug administration. The skin irritation score ranged from 0 to 0.33, indicating no skin irritation. The irritation response scores for single application to the damaged skin of rabbits are shown in Table 18, and the irritation response scores for multiple application to the damaged skin of rabbits are shown in Table 19.

[0261] Table 16. Irritation response scores of rabbits after a single application of HQBG to intact skin (n=6)

[0262]

[0263] Table 17. Irritation response scores of rabbits after repeated application of HQBG to intact skin (n=6)

[0264]

[0265]

[0266] Table 18 shows the irritation response scores of rabbits after a single application of HQBG to broken skin (n=6).

[0267]

[0268] Table 19 shows the irritation response scores of rabbits after repeated application of HQBG to damaged skin (n=6).

[0269]

[0270]

[0271] Although the present invention has been described in detail above with general descriptions and specific embodiments, modifications or improvements can be made to it, which will be obvious to those skilled in the art. Therefore, all such modifications or improvements made without departing from the spirit of the present invention fall within the scope of protection claimed by the present invention.

Claims

1. A traditional Chinese medicine composition for treating eczema, comprising the following components: 1 part by weight of indigo naturalis, 1.6-2.4 parts by weight of gypsum, 1.2-1.8 parts by weight of phellodendron bark, 2.4-3.6 parts by weight of talc, 1.04-1.56 parts by weight of isatis leaf, 1.12-1.68 parts by weight of alisma rhizome, 1.36-2.04 parts by weight of poria cocos, 1.12-1.68 parts by weight of kochia fruit, 0.8-1.2 parts by weight of licorice root, and 0.96-1.44 parts by weight of sanguisorba officinalis root.

2. A traditional Chinese medicine extract for treating eczema, obtained by alcohol extraction after mixing various raw materials; the raw materials are as follows: 1 part by weight of indigo naturalis, 1.6-2.4 parts by weight of gypsum, 1.2-1.8 parts by weight of phellodendron bark, 2.4-3.6 parts by weight of talc, 1.04-1.56 parts by weight of isatis leaf, 1.12-1.68 parts by weight of alisma rhizome, 1.36-2.04 parts by weight of poria cocos, 1.12-1.68 parts by weight of kochia fruit, 0.8-1.2 parts by weight of licorice root, and 0.96-1.44 parts by weight of sanguisorba officinalis root; The extraction reagent used for the alcohol extraction is ethanol with a volume fraction of 60-80%.

3. The herbal extract according to claim 2, characterized in that: The conditions for alcohol extraction are: reflux in a water bath at a temperature of 75-90℃ for 0.5-2 hours.

4. A traditional Chinese medicine preparation comprising the traditional Chinese medicine composition of claim 1 or the traditional Chinese medicine extract of claim 2 or 3.

5. The traditional Chinese medicine preparation according to claim 4, characterized in that: The dosage forms of the traditional Chinese medicine preparations are gels, liposomes, creams, and ointments.

6. The traditional Chinese medicine preparation according to claim 4, characterized in that: The dosage form of the traditional Chinese medicine preparation is a microemulsion gel.

7. The traditional Chinese medicine preparation according to claim 5, characterized in that: The gelling agent comprises the following components: The traditional Chinese medicine composition according to claim 1 or the traditional Chinese medicine extract, matrix and adjuvants according to claim 2 or 3; In the gel, the mass fraction of the traditional Chinese medicine composition or the traditional Chinese medicine extract is 3-7%; The mass fraction of the matrix is ​​0.8-1.2%; The mass fraction of the auxiliary agent is 11.55-22.2%.

8. The traditional Chinese medicine preparation according to claim 7, characterized in that: The matrix is ​​one or more of carbomer, chitosan, poloxamer, and cellulose; The additives include humectants, pH adjusters, and preservatives.

9. The traditional Chinese medicine preparation according to claim 8, characterized in that: The moisturizer is glycerin or propylene glycol; The pH adjuster is triethanolamine or sodium hydroxide; The preservative is one or more of potassium sorbate, methylparaben, and ethylparaben.

10. The traditional Chinese medicine preparation according to claim 9, characterized in that: The mass fraction of the humectant in the gel is 10-20%; The mass fraction of the pH adjuster in the gel is 1.5-2%; The preservative in the gel has a mass fraction of 0.05-0.2%.

11. The use of the traditional Chinese medicine composition according to claim 1, the traditional Chinese medicine extract according to claim 2 or 3, or the traditional Chinese medicine preparation according to any one of claims 4-10 in the preparation of a drug for treating eczema.

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