A composition of active substances from traditional Chinese medicine with anti-psoriasis effects, its preparation method and application in cream form.

Abstract

This invention discloses a composition of traditional Chinese medicine active substances with anti-psoriasis effects, its preparation method for an ointment, and its application. The composition consists of baicalin, the main active ingredient of Scutellaria baicalensis, and echinacoside, the main active ingredient of Cistanche deserticola. Baicalin can inhibit skin inflammatory infiltration through the TNF signaling pathway, while echinacoside can inhibit skin angiogenesis through the VEGF signaling pathway, thereby inhibiting the development of psoriatic lesions. This invention, through pharmacological experiments, screened and obtained an optimal combination of baicalin and echinacoside. This composition can significantly inhibit the pathological progression of psoriatic lesions by synergistically regulating multiple pathways, multiple targets, and multiple processes, and has good clinical application value.

Description

Technical Field

[0001] This invention relates to a composition of active substances from traditional Chinese medicine and its preparation method and uses, and particularly to a composition of active substances from traditional Chinese medicine with anti-psoriasis effects and its preparation method and applications. Background Technology

[0002] Psoriasis, commonly known as "cowhide rash," is a chronic inflammatory skin disease characterized by excessive proliferation of epidermal keratinocytes, dermal microvascular proliferation, and inflammatory cell infiltration. It is a multifactorial, multi-stage disease that clinically manifests as skin lesions characterized by erythema, scaling, and inflammatory thickening. Based on different clinical phenotypes, psoriasis is classified into plaque psoriasis, pustular psoriasis, psoriatic arthritis, and erythrodermic psoriasis, with plaque psoriasis being the most common.

[0003] Currently, topical medications for psoriasis lesions mainly consist of retinoids, vitamin D3 analogs, and corticosteroids. Retinoids and vitamin D3 analogs can inhibit excessive proliferation of epidermal keratinocytes, while corticosteroids can suppress excessive inflammatory responses. However, these topical medications all have varying degrees of side effects, such as skin irritation and contraindications for long-term use. In particular, long-term use of corticosteroids can cause skin atrophy and telangiectasia. Therefore, developing a non-steroidal topical medication that is well-adapted to the skin, has few side effects, significant efficacy, can act on multiple stages of the process, and can be used long-term is of paramount importance.

[0004] Traditional Chinese medicine and its active substances have the characteristics of relatively low toxicity and few side effects, making them suitable for long-term use, and they show broad application prospects, especially in the treatment of chronic inflammatory diseases. Summary of the Invention

[0005] Purpose of the invention: The purpose of this invention is to provide a composition of active substances from traditional Chinese medicine that has anti-psoriasis effects.

[0006] Another object of the present invention is to provide a method for preparing an ointment of the composition of traditional Chinese medicine active substances with anti-psoriasis effects.

[0007] Another object of the present invention is to provide the use of the composition of active substances of traditional Chinese medicine with anti-psoriasis effects.

[0008] This invention targets the external skin lesions of psoriasis, aiming to develop a topical medication with good skin adaptability, few side effects, significant efficacy, and the ability to act on multiple stages of the disease process for long-term use. The composition is obtained by combining baicalin, the main active ingredient of Scutellaria baicalensis, and echinacoside, the main active ingredient of Cistanche deserticola. Pharmacological studies have shown that these two active ingredients can respectively regulate different pathological processes in the development of psoriatic skin lesions (inhibiting inflammatory infiltration and inhibiting angiogenesis), thereby exerting a synergistic effect in improving psoriatic skin lesions.

[0009] Baicalin is the main active ingredient of the traditional Chinese medicine Scutellaria baicalensis, and it has significant anti-inflammatory, anti-allergic, and antioxidant effects. Echinacoside is the main active ingredient of the traditional Chinese medicine Cistanche deserticola, and also the main active ingredient of the natural medicinal plant Echinacea purpurea (also known as purple echinacea or coneflower), and it has significant antioxidant, anti-angiogenic, and anti-inflammatory effects.

[0010] The combination of active ingredients in traditional Chinese medicine can exert a synergistic effect. However, there are currently no research reports on the synergistic regulation of multiple pathological processes in the development of psoriatic lesions through the combination of baicalin, the main active ingredient of Scutellaria baicalensis, and echinacoside, the main active ingredient of Cistanche deserticola.

[0011] Specifically,

[0012] (1) The baicalin and echinacoside used in this invention are natural active ingredients derived from medicinal plants. Existing studies have confirmed that they have significant pharmacological effects and are essentially non-toxic to cells. Compared with glucocorticoid drugs, retinoid drugs, and vitamin D3 analog drugs, these two natural plant active ingredients are safer and have almost no skin irritation.

[0013] (2) Based on the physicochemical properties of baicalin and echinacoside, this invention uses cream-type matrix excipients and employs a multi-factor orthogonal experimental method to investigate factors such as emulsifier dosage, emulsification temperature, and emulsification time, thereby optimizing the best preparation method for the cream. Baicalin and echinacoside are stable and do not precipitate in this cream, resulting in a uniform, fine, and easily spreadable cream.

[0014] (3) This invention utilizes a stable and usable composition cream. First, its efficacy was observed in an imiquimod (IMQ)-induced mouse psoriasis model, revealing that the composition significantly improved the skin lesion phenotype. Then, network pharmacology and bioinformatics were used to predict that baicalin might combat psoriasis through the TNF signaling pathway, and echinacoside might combat psoriasis through the VEGF signaling pathway. Further pharmacological experiments were conducted to investigate and verify these findings. Baicalin significantly inhibited the release of inflammatory factors such as TNF-α and suppressed inflammatory cell infiltration, while echinacoside significantly inhibited VEGF-A expression, suppressed angiogenesis, and reduced oxidative damage. Finally, through the detection of various experimental indicators, the optimal composition ratio was obtained. This composition ratio exhibited more significant pharmacological effects than single-component compositions and could effectively synergistically regulate multiple pathological stages in the development of psoriatic skin lesions.

[0015] Technical solution: The active ingredient composition of traditional Chinese medicine with anti-psoriasis effect provided by the present invention includes the following raw materials in parts by weight: 1-10 parts by weight of baicalin and 1-10 parts by weight of echinacoside.

[0016]

[0017] Preferably, the raw materials include the following parts by weight: 1-2 parts by weight of baicalin and 1-2 parts by weight of echinacoside.

[0018] A pharmaceutical preparation comprising the aforementioned composition of active traditional Chinese medicine substances with anti-psoriasis effects, wherein the preparation is for external use.

[0019] The above-mentioned pharmaceutical preparation is a cream for external use.

[0020] The above-described cream is prepared from a composition of base excipients and active traditional Chinese medicine substances in the following weight percentages:

[0021] Oil phase:

[0022] Stearic acid 8-12%

[0023] Glyceryl monostearate 4-8%

[0024] 3-4% white petrolatum

[0025] Aqueous phase:

[0026]

[0027] Ultrapure water balance

[0028] The preparation method of the cream includes the following steps:

[0029] (1) Heat 8-12% stearic acid, 4-8% glyceryl monostearate and 3-4% white petrolatum in a water bath to 78-82°C to melt them into liquid and form an oil phase;

[0030] (2) 4-6% triethanolamine, 4-8% glycerol, 1-3% azone, 4-8% of the composition, 0.05-0.2% ethylparaben, and a portion of ultrapure water are heated in a water bath to 78-82°C to completely dissolve them into a solution, forming an aqueous phase;

[0031] (3) While the oil phase is hot, add the remaining ultrapure water and stir continuously until it is evenly mixed. After cooling to 40°C, continue stirring until it is completely emulsified. After cooling to room temperature, the cream is obtained.

[0032] Beneficial effects: Compared with the prior art, the present invention has the following advantages:

[0033] The baicalin in the formulation of this invention can inhibit skin inflammatory infiltration through the TNF signaling pathway, and echinacoside can inhibit skin angiogenesis through the VEGF signaling pathway, thereby inhibiting the development of psoriatic lesions. This invention has screened the optimal ratio of baicalin and echinacoside through pharmacological experiments to obtain a suitable combination. This combination can significantly inhibit the pathological progression of psoriatic lesions by synergistically regulating multiple pathways, multiple targets, and multiple processes, and has good clinical application value. Attached Figure Description

[0034] Figure 1 This section shows the changes in dorsal skin lesions in each group of mice in Example 3 (n=8); A shows the appearance of the dorsal skin lesions in each group of mice on the last day of the experiment; B shows the dorsal skin lesion indices (erythema score, scales score, thickness score, and cumulative score) in each group of mice during the experiment. Changes in Score; Con group: normal control group, IMQ group: imiquimod (IMQ) induced psoriasis model group, KB group: IMQ induced + blank cream matrix group, HMS group: IMQ induced + halometasone cream positive control group, BAI group: IMQ induced + 5% baicalin cream administration group, ECH group: IMQ induced + 5% echinacoside cream administration group, B1-E1 group: IMQ induced + 5% composition 1 (baicalin: echinacoside = 1:1) cream administration group, B2-E1 group: IMQ induced + 5% composition 2 (baicalin: echinacoside = 2:1) cream administration group, B1-E2 group: IMQ induced + 5% composition 3 (baicalin: echinacoside = 1:2) cream administration group;

[0035] Figure 2HE staining results of dorsal skin lesions in each group of mice in Example 3 (n=6); double arrows indicate epidermal thickness; red circles indicate inflammatory cell infiltration;

[0036] Figure 3 The network pharmacology prediction results of the potential mechanism of action of baicalin against psoriasis in Example 3 are shown in A; the network pharmacology prediction shows the potential mechanism of action of baicalin against psoriasis; and the virtual docking analysis of baicalin with TNF, a key target of the TNF signaling pathway, is shown in B.

[0037] Figure 4 The network pharmacology prediction results of the potential mechanism of action of echinacoside against psoriasis in Example 3 are shown in A; the network pharmacology prediction shows the potential mechanism of action of echinacoside against psoriasis; and the virtual docking analysis of echinacoside with VEGFA, a key target of the VEGF signaling pathway, is shown in B.

[0038] Figure 5 The immunohistochemical staining results of CD68, a marker of macrophage inflammatory infiltration, in the dorsal skin lesions of mice in each group in Example 3 (n=6);

[0039] Figure 6 The effects of each experimental group of drugs on angiogenesis in zebrafish larvae in Example 4 (n=6); CON group: normal control group, VRI group: positive control group for angiogenesis loss in zebrafish larvae induced by vascular endothelial growth factor receptor tyrosine kinase inhibitor II (VRI), BAI group: 50 μg / mL baicalin administration group, ECH group: 50 μg / mL echinacoside administration group, B1-E1 group: 50 μg / mL composition 1 (baicalin:echinacoside = 1:1) administration group, B2-E1 group: 50 μg / mL composition 2 (baicalin:echinacoside = 2:1) administration group, B1-E2 group: 50 μg / mL composition 3 (baicalin:echinacoside = 1:2) administration group. Detailed Implementation

[0040] Example 1

[0041] A topical cream with anti-psoriasis effects, comprising the following components (by weight percentage):

[0042] Oil phase:

[0043] Stearic acid 9.5%

[0044] 5% glyceryl monostearate

[0045] 4% White Vaseline

[0046] Aqueous phase:

[0047]

[0048] Ultrapure water balance

[0049] The preparation method of the above-mentioned topical cream with anti-psoriasis effects is as follows:

[0050] (1) Heat 9.5% stearic acid, 5% glyceryl monostearate and 4% white petrolatum in a water bath to 80°C to melt them into liquid and form an oil phase;

[0051] (2) 5% triethanolamine, 5.5% glycerol, 2% azone, 5% active traditional Chinese medicine composition, 0.09% ethylparaben, and a portion of ultrapure water are heated to 80°C in a water bath to completely dissolve them into a solution, forming an aqueous phase;

[0052] (3) While the oil phase is hot, add the remaining ultrapure water and stir continuously for 15 minutes. After cooling to 40°C, continue stirring for 15 minutes until completely emulsified. After cooling to room temperature, the cream is obtained.

[0053] (4) The cream is packaged into 30g boxes.

[0054] Example 2

[0055] The optimal preparation process of the cream was selected using a multi-factor orthogonal experimental method.

[0056] 1. Instruments and Materials

[0057] 1.1 Instruments

[0058] METTLER TOLEDO AL204 electronic balance (Mettler-Toledo Instruments (Shanghai) Co., Ltd.); Milli-Q ultrapure water system (Millipore, USA); JOANLAB water bath (Ningbo Qun'an Experimental Instruments Co., Ltd.); Eppendorf 5804R high-speed refrigerated centrifuge (Eppendorf, Germany); electric heating drying oven (Shanghai Yiheng Scientific Instruments Co., Ltd.).

[0059] 1.2 Materials

[0060] Baicalin (HPLC purity ≥98%, Baoji Chenguang Biotechnology Co., Ltd.); Echinacoside (HPLC purity ≥94%, Baoji Chenguang Biotechnology Co., Ltd.); Stearic acid (analytical grade, Sinopharm Chemical Reagent Co., Ltd.); White petrolatum (medical grade, Shanghai Aladdin Biochemical Technology Co., Ltd.); Glyceryl monostearate (emulsified type, purity 99%, Shanghai Aladdin Biochemical Technology Co., Ltd.); Ethylparaben (analytical grade, Shanghai Aladdin Biochemical Technology Co., Ltd.); Triethanolamine (analytical grade, Shanghai Aladdin Biochemical Technology Co., Ltd.); Glycerol (analytical grade, Sinopharm Chemical Reagent Co., Ltd.); Azone (pharmaceutical grade, Shanghai Aladdin Biochemical Technology Co., Ltd.).

[0061] 2. Methods and Results

[0062] 2.1 Prescription and Preparation

[0063] The formulation and preparation method of the cream are described in Example 1.

[0064] 2.2 Cream Scoring Criteria

[0065] The total score is set at 100 points, with 20 points each for appearance quality, pH, centrifugal stability, and heat and cold resistance. See Table 1 for specific scoring criteria.

[0066] Table 1. Cream Scoring Criteria

[0067]

[0068]

[0069] 2.3 Optimization of Preparation Process

[0070] The contents of triethanolamine (Factor A) and glyceryl monostearate (Factor B), emulsification temperature (Factor C), and emulsification time (Factor D) were used as the factors to be investigated, and an orthogonal experimental factor level table was designed (see Table 2). The comprehensive score was used as the evaluation index, and an L9(3)0 ... 4 The orthogonal experimental design preparation process is shown in Table 3, and SPSS 19.0 statistical software was used for analysis. The results of the orthogonal experimental variance analysis are shown in Table 4.

[0071] Table 3 shows that the order of influence of each factor on the experimental results is D > C > A > B, and the optimal preparation process is A3B2C2D1, namely, the amount of triethanolamine is 5%, the amount of glyceryl monostearate is 5%, the emulsification temperature is 80℃, and the emulsification time is 15min. Table 4 shows that each factor has a significant impact on the experimental results. Therefore, this condition is determined as the optimal preparation process for the cream, and based on this, 5% baicalin cream, 5% echinacoside cream, 5% composition 1 (baicalin:echinacoside = 1:1) cream, 5% composition 2 (baicalin:echinacoside = 2:1) cream, and 5% composition 3 (baicalin:echinacoside = 1:2) cream are prepared respectively.

[0072] Table 2. Factor Level Table for Orthogonal Experiment

[0073]

[0074] Table 3 L9(3) 4 Orthogonal experimental design scheme and results

[0075]

[0076]

[0077] Table 4 L9(3 4 ) Orthogonal experiment variance analysis results

[0078]

[0079] Example 3

[0080] Study on the pharmacodynamic effects and mechanisms of creams with different drug ratios prepared in Example 2 against psoriasis

[0081] 1. Instruments and materials

[0082] 1.1 Instruments

[0083] METTLER TOLEDO AL204 electronic balance (METTLER TOLEDO Instruments (Shanghai) Co., Ltd.); Milli-Q ultrapure water instrument (Millipore, USA); JOANLAB water bath (Ningbo Qunan Experimental Instrument Co., Ltd.); OLYMPUS BX53 upright fluorescence microscope (OLYMPUS, Japan); high-throughput cryogenic grinder (Nanjing Zhongke); Sigma 2-16K high-speed refrigerated centrifuge (Sigma, Germany); TECAN Sunrise microplate reader (TECAN, Switzerland). [[ID=2)]]

[0084] 1.2 Materials

[0085] The experimental animals were 8-week-old female BALB / c mice, SPF grade, weighing 16 - 19 g, 72 in number, purchased from Zhejiang Vital River Laboratory Animal Technology Co., Ltd., with the certificate number 20220602Abzz0619000465 and the production license number SCXK(Zhe)2019-0001, and were housed in the Experimental Animal Center of the Institute of Dermatology, Chinese Academy of Medical Sciences.

[0086] Homemade 5% baicalin cream, 5% echinacoside cream, 5% Composition 1 (baicalin:echinacoside = 1:1) cream, 5% Composition 2 (baicalin:echinacoside = 2:1) cream, 5% Composition 3 (baicalin:echinacoside = 1:2) cream, and blank cream (the blank cream contains no drugs, and the proportions of other matrix excipients remain unchanged); imiquimod cream (Adal brand, Inova Pharmaceutical); halometasone cream (Aoneng brand, Aomei Pharmaceutical); tissue fixative (Jiangsu Shitai Pharmaceutical). Test Equipment Co., Ltd.); Mouse Tumor Necrosis Factor-α (TNF-α), Interleukin-17A (IL-17A), Interleukin-23 (IL-23), Interleukin-10 (IL-10) ELISA kits and Vascular Endothelial Growth Factor A (VEGF-A), Matrix Metalloproteinase-9 (MMP-9), Thioredoxin Reductase (TrxR), Malondialdehyde (MDA) ELISA kits (Nanjing Senbeijia Biotechnology Co., Ltd.); Anti-CD68 Rabbit pAb (Wuhan Saiweier Biotechnology Co., Ltd.).

[0087] 2. Methods

[0088] 2.1 Preparation of mouse psoriasis model and drug intervention

[0089] Imiquimod (IMQ)-induced mouse psoriasis models reproduce the typical clinical symptoms and pathological features of human psoriasis, such as skin erythema, scaling, epidermal thickening, and inflammatory infiltration. Therefore, this invention uses the widely used IMQ-induced mouse psoriasis model to study the efficacy and mechanism of creams with different drug ratios against psoriasis.

[0090] Seventy-two mice were randomly divided into nine groups: (1) Con group: normal control group; (2) IMQ group: imiquimod (IMQ) induced psoriasis model group; (3) KB group: IMQ induced + blank cream matrix group; (4) HMS group: IMQ induced + halometasone cream positive control group; (5) BAI group: IMQ induced + 5% baicalin cream administration group; (6) ECH group: IMQ induced + 5% echinacoside cream administration group; (7) B1-E1 group: IMQ induced + 5% composition 1 (baicalin: echinacoside = 1:1) cream administration group; (8) B2-E1 group: IMQ induced + 5% composition 2 (baicalin: echinacoside = 2:1) cream administration group; (9) B1-E2 group: IMQ induced + 5% composition 3 (baicalin: echinacoside = 1:2) cream administration group. Except for the Con group, mice in all other groups had 62.5 mg of 5% imiquimod cream applied daily to the hairless (3 days before formal modeling) dorsal skin (2cm×3cm) for 6 consecutive days to establish a mouse psoriasis model. Except for the IMQ group, all cream groups had the cream applied 4 hours after the IMQ application, with the same dosage as the imiquimod cream.

[0091] Modeling and drug administration began on day 0 and continued until day 5, with skin lesions scored daily. On day 6, mice in each group were photographed and scored, and then skin tissue was harvested from the backs of the mice. A portion was preserved in tissue fixative for HE staining and immunohistochemical staining, while the other portion was preserved at -80℃ for biochemical index testing.

[0092] 2.2 Skin lesion score

[0093] The severity of back skin lesions was assessed using the Psoriasis Area and Severity Index (PASI) scoring system, which assesses the severity of erythema, scaling, and infiltration thickness. Each parameter was scored independently on a scale of 0 to 4, where 0 = asymptomatic, 1 = mild symptoms, 2 = moderate symptoms, 3 = significant symptoms, and 4 = very significant symptoms. The cumulative score represents the severity of inflammation.

[0094] 2.3 HE staining

[0095] Skin tissue samples from mice in each group were stained with hematoxylin and eosin (HE) to observe epidermal thickening and inflammatory infiltration.

[0096] 2.4 Network Pharmacology Prediction

[0097] Downloaded SDF files of baicalin and echinacoside from the PubChem database and converted them to SMILE and MO2 formats using OpenBabel 2.4.1. The MO2 format files were uploaded to the PharmMapper database, and the Human Protein Targets Only option was selected to predict the top 300 target molecules with the highest matching degree. Then, using the SMILE format files of baicalin and echinacoside, the Homo sapiens option was selected to predict the target molecules of baicalin and echinacoside in the SEA and SIB databases, respectively. Finally, the predicted target molecules of baicalin and echinacoside were summarized. Psoriasis disease targets were searched in the GeneCards, DrugBank, and DisGeNET databases. The predicted target molecules of baicalin and echinacoside were mapped to psoriasis disease targets to screen for potential anti-psoriasis targets of baicalin and echinacoside. The interaction relationships between targets were further analyzed using the STRING database, and the KOBAS database was further used to perform KEGG pathway enrichment analysis on key targets, and virtual docking analysis was performed on key targets.

[0098] 2.5 Biochemical index detection

[0099] Skin tissue samples were collected, subcutaneous fat was removed, and the samples were weighed, minced, and added to pre-cooled PBS in a ratio of 1:9 (tissue weight:PBS volume = 1:9) and homogenized. The homogenate was centrifuged at 3000 rpm for 15 min at 4°C, and the supernatant was collected. The protein concentration of each group of samples was determined using the BCA method. According to the ELISA kit instructions, the contents of tumor necrosis factor-α (TNF-α), interleukin-17A (IL-17A), interleukin-23 (IL-23), interleukin-10 (IL-10), vascular endothelial growth factor A (VEGF-A), matrix metalloproteinase-9 (MMP-9), thioredoxin reductase (TrxR), and malondialdehyde (MDA) in the skin lesions were determined.

[0100] 2.6 Immunohistochemical staining

[0101] Immunohistochemical staining was performed on skin tissue samples from each group of mice to observe the positive expression of CD68, a marker of macrophage inflammatory infiltration.

[0102] 2.7 Statistical Analysis

[0103] Experimental data are expressed as mean ± standard deviation. One-way ANOVA was used for comparisons between multiple groups, and Sidak's multiple comparisons were used for pairwise comparisons. Data were analyzed using GraphPad Prism 8 software, and P < 0.05 was defined as statistically significant.

[0104] 3. Results

[0105] 3.1 Effects of creams with different drug ratios on IMQ-induced psoriatic lesions in mice

[0106] The severity of skin lesions on the backs of mice was assessed using the Psoriasis Lesion Severity Index, and the results were as follows: Figure 1 As shown, compared with the Con group, mice in the IMQ and KB groups exhibited severe skin lesions on their backs. Compared with the IMQ and KB groups, creams prepared with different ratios of baicalin and echinacoside significantly reduced skin erythema, scaling, and thickening, thus decreasing the severity of skin inflammation. HE staining and morphometric analysis of the skin lesions ( Figure 2 Table 5 shows that creams prepared with different ratios of baicalin and echinacoside significantly improved IMQ-induced epidermal thickening (P<0.001) and reduced inflammatory cell infiltration after application. Among them, the cream with 5% composition 2 (baicalin:echinacoside = 2:1) showed more significant effects and was superior to single-component baicalin and echinacoside to a certain extent.

[0107] Table 5. Epidermal thickness of dorsal skin tissue in mice of each group

[0108]

[0109] Note: Data are expressed as mean ± standard deviation (n = 6). Compared with the Con group, ΔΔΔ P<0.001; compared with the IMQ group, ### P<0.001; compared with the KB group, ***P <0.001.

[0110] 3.2 Prediction of the potential mechanisms of action of baicalin and echinacoside in the treatment of psoriasis

[0111] like Figure 3 As shown, the potential key target of baicalin in anti-psoriasis is significantly enriched in the TNF signaling pathway, and molecular docking shows that baicalin has a good affinity for TNF, a key target in the TNF signaling pathway, indicating that baicalin may exert its anti-psoriasis effect by regulating the TNF signaling pathway.

[0112] like Figure 4As shown, the potential key target of echinacoside in anti-psoriasis is significantly enriched in the VEGF signaling pathway, and molecular docking shows that echinacoside has a strong affinity for VEGFA, a key target in the VEGF signaling pathway, indicating that echinacoside may exert its anti-psoriasis effect by regulating the VEGF signaling pathway.

[0113] 3.3 Effects of creams with different drug ratios on the release of inflammatory factors and infiltration of inflammatory cells in psoriatic lesions of mice

[0114] Figure 3 The key target TNF in the TNF signaling pathway predicted by the network pharmacology is TNF, namely tumor necrosis factor-α (TNF-α). TNF-α can mediate inflammatory responses, involving the release of inflammatory factors and the infiltration of inflammatory cells. As shown in Table 6, compared with the Con group, the levels of inflammatory factors TNF-α, IL-17A, and IL-23 in the skin lesions of mice in the IMQ and KB groups were significantly increased (P<0.001), while the levels of anti-inflammatory factor IL-10 were significantly decreased (P<0.001). Compared with the IMQ and KB groups, the creams prepared with different ratios of baicalin and echinacoside could reduce the levels of inflammatory factors and increase the content of anti-inflammatory factors to varying degrees (P<0.05, P<0.01, P<0.001). The 5% composition 2 (baicalin:echinacoside = 2:1) cream had a more significant effect and was superior to the single components of baicalin and echinacoside to a certain extent.

[0115] TNF-α-mediated inflammatory responses also involve the infiltration of inflammatory cells such as macrophages. Immunohistochemical staining results of the macrophage marker CD68 in skin lesions (…). Figure 5 The results showed that, compared with the Con group, the positive expression of the macrophage marker CD68 in the skin lesions of mice in the IMQ and KB groups was significantly increased (the brown areas are positive staining areas of CD68); compared with the IMQ and KB groups, the creams prepared with different ratios of baicalin and echinacoside could reduce the positive expression of CD68 to varying degrees, indicating that the inflammatory infiltration of macrophages was inhibited, and the 5% composition 2 (baicalin:echinacoside = 2:1) cream also showed a more significant effect.

[0116] Table 6. Content of inflammatory and anti-inflammatory factors in the skin lesions of mice in each group.

[0117]

[0118]

[0119] Note: Data are expressed as mean ± standard deviation (n = 6). Compared with the Con group, ΔΔΔ P<0.001; compared with the IMQ group, # P<0.05, ##P<0.01, ### P<0.001; compared with the KB group, * P<0.05, ** P<0.01, *** P<0.001.

[0120] 3.4 Effects of creams with different drug ratios on vascular endothelial growth factor expression and oxidative stress response in psoriatic lesions of mice

[0121] Figure 4 The network pharmacology predicted a key target in the VEGF signaling pathway, namely vascular endothelial growth factor A (VEGF-A). VEGF-A is closely related to the regulation of angiogenesis, which is also closely related to another key factor, matrix metalloproteinase-9 (MMP-9). MMP-9 can participate in angiogenesis by releasing VEGF. As shown in Table 7, compared with the Con group, the levels of angiogenesis-related factors VEGF-A and MMP-9 in the skin lesions of mice in the IMQ and KB groups were significantly increased (P<0.001), indicating the presence of angiogenesis. Compared with the IMQ and KB groups, the creams prepared with different ratios of baicalin and echinacoside could reduce the levels of VEGF-A and MMP-9 to varying degrees (P<0.01, P<0.001). The 5% composition 2 (baicalin:echinacoside = 2:1) cream had a more significant effect and was superior to the single components of baicalin and echinacoside to a certain extent.

[0122] Studies have shown that the regulation of angiogenesis is also closely related to antioxidant activity. As shown in Table 7, compared with the Con group, the activity of TrxR, an indicator of oxidative stress, was significantly reduced and the content of MDA was significantly increased in the skin lesions of mice in the IMQ and KB groups (P<0.001), indicating oxidative damage. Compared with the IMQ and KB groups, the creams prepared with different ratios of baicalin and echinacoside could increase TrxR activity and decrease MDA content to varying degrees (P<0.01, P<0.001), indicating enhanced antioxidant activity. The 5% composition 2 (baicalin:echinacoside = 2:1) cream had a more significant effect.

[0123] Table 7. Levels of angiogenesis-related factors and antioxidant capacity in skin lesions of mice in each group.

[0124]

[0125] Note: Data are expressed as mean ± standard deviation (n = 6). Compared with the Con group, ΔΔΔ P<0.001; compared with the IMQ group, ## P<0.01, ### P<0.001; compared with the KB group, *** P<0.001.

[0126] Example 4

[0127] Study on the anti-angiogenic effects and mechanisms of baicalin and echinacoside in different ratios

[0128] 1. Instruments and Materials

[0129] 1.1 Instruments

[0130] METTLER TOLEDO AL204 electronic balance (Mettler-Toledo Instruments (Shanghai) Co., Ltd.); Milli-Q ultrapure water system (Millipore, USA); KH-500B ultrasonic cleaner (Kunshan Hechuang Ultrasonic Instruments Co., Ltd.); SHP-250 biochemical incubator (Shanghai Senxin Experimental Instruments Co., Ltd.); OLYMPUS BX53 upright fluorescence microscope (OLYMPUS, Japan); Lightcycler 480 fully automated real-time PCR instrument (Roche, USA).

[0131] 1.2 Materials

[0132] Baicalin (HPLC purity ≥98%, Baoji Chenguang Biotechnology Co., Ltd.); Echinacoside (HPLC purity ≥94%, Baoji Chenguang Biotechnology Co., Ltd.); DMSO (analytical grade, Shanghai Titan Technology Co., Ltd.); Streptomycin (biological reagent, 7000u / g, Shanghai Yuanye Biotechnology Co., Ltd.); Vascular endothelial growth factor receptor tyrosine kinase inhibitor II (VRI) (HPLC purity >99.0%, Merck, Germany); RNA extraction reagent, reverse transcription kit, qPCR kit (Hunan Aikerui Biotechnology Co., Ltd.); PCR primers were synthesized by Sangon Biotech (Shanghai) Co., Ltd.; Tg(fli-1:EGFP)y1 transgenic zebrafish embryos (vascular endothelial cell-specific green fluorescence) and embryo culture medium were purchased from Nanjing Yishu Lihua Biotechnology Co., Ltd.

[0133] 2. Methods

[0134] 2.1 Preparation of drug solution

[0135] Weigh an appropriate amount of the drug to be used in the experiment, dissolve it in an appropriate amount of DMSO, dilute it with zebrafish embryo culture medium, and bring the volume to a suitable level to obtain a drug stock solution. Before use, take an appropriate amount of the stock solution and dilute it with culture medium to the required concentration.

[0136] 2.2 Inhibition of angiogenesis in Tg(fli-1:EGFP)y1 transgenic zebrafish larvae

[0137] Zebrafish embryos in good condition and developed to 24 hours were selected. The eggshells were removed with 1 mg / mL streptomycin solution, and the embryos were placed in 24-well plates with at least 10 embryos per well as one group. The CON group was given zebrafish embryo culture medium containing 0.1% DMSO, the VRI group was given 500 ng / mL VRI, the BAI group was given 50 μg / mL baicalin, the ECH group was given 50 μg / mL echinacoside, the B1-E1 group was given 50 μg / mL composition 1 (baicalin:echinacoside = 1:1), the B2-E1 group was given 50 μg / mL composition 2 (baicalin:echinacoside = 2:1), and the B1-E2 group was given 50 μg / mL composition 3 (baicalin:echinacoside = 1:2). After incubating at 28℃ for 24 hours, zebrafish juveniles were placed on biconcave slides and kept in a lateral position. The growth of intersegmental vessels (ISVs) in each group of zebrafish juveniles was observed using an upright fluorescence microscope. The number of intact and missing intersegmental vessels in each group was counted, and the ISV index was calculated. ISV index = (number of intact intersegmental vessels × 1) + (number of missing intersegmental vessels × 0.5).

[0138] 2.3 qPCR detection of angiogenesis-related gene expression in VEGF signaling pathway in zebrafish juveniles

[0139] The initial zebrafish embryo treatment and drug treatment procedures were the same as described in section 2.2. After incubation at 28°C for 24 hours, total RNA was extracted from each group of zebrafish larvae using RNA extraction reagent. The total RNA was then reverse transcribed into cDNA using a reverse transcription kit, followed by qPCR experiments. β-actin was used as an internal reference gene to detect the expression of angiogenesis-related genes VEGF-A, kdrl, flt-1, and vWF in zebrafish larvae.

[0140] 2.4 Statistical Analysis

[0141] Experimental data are expressed as mean ± standard deviation. One-way ANOVA was used for comparisons between multiple groups, and Dunnett's multiple comparisons were used for pairwise comparisons. Data were analyzed using GraphPad Prism 8 software, and P < 0.05 was defined as statistically significant.

[0142] 3. Results

[0143] 3.1 Effects of different ratios of baicalin and echinacoside on intersegmental angiogenesis in juvenile zebrafish

[0144] like Figure 6As shown, the intersegmental vessels of zebrafish juveniles in the normal control group (CON group) remained intact, and angiogenesis was not affected. Intersegmental angiogenesis in the VRI group juveniles was significantly inhibited, and intersegmental angiogenesis in the BAI, ECH, B1-E1, B2-E1, and B1-E2 groups was also inhibited to varying degrees. The intersegmental vessel index of each group of zebrafish juveniles was calculated by statistically analyzing the number of intact and defective intersegmental vessels (see Table 8). The results showed that compared with the CON group, the BAI, ECH, B1-E1, B2-E1, and B1-E2 groups all significantly inhibited intersegmental angiogenesis in zebrafish juveniles (P<0.001). Among the compositions, the B2-E1 group (baicalin:echinacea glycoside = 2:1) showed a more significant effect, significantly superior to the single components of baicalin and echinacea glycoside.

[0145] Table 8. Intersegmental vascular index of zebrafish juveniles in each group.

[0146]

[0147] Note: Data are expressed as mean ± standard deviation (n = 6). Compared with the CON group, *** P<0.001.

[0148] 3.2 Effects of different ratios of baicalin and echinacoside on the expression of angiogenesis-related genes in the VEGF signaling pathway of zebrafish juveniles

[0149] As shown in Table 9, qPCR results indicated that the expression of angiogenesis-related genes VEGF-A, kdrl, flt-1, and vWF in zebrafish juveniles in the VRI positive control group was significantly decreased compared to the CON group (P<0.001). Compared to the CON group, the expression of angiogenesis-related genes in zebrafish juveniles in the BAI, ECH, B1-E1, B2-E1, and B1-E2 groups was also inhibited to varying degrees (P<0.05, P<0.01, P<0.001). Among the compositions, the B2-E1 group (baicalin:echinacea glycoside = 2:1) showed a more significant effect, significantly superior to the single components of baicalin and echinacea glycoside. These studies suggest that the mechanism of action of the baicalin and echinacea glycoside combination in inhibiting angiogenesis may be related to the inhibition of the expression of angiogenesis-related genes VEGF-A, kdrl, flt-1, and vWF in the VEGF signaling pathway.

[0150] Table 9. Expression of genes related to angiogenesis in zebrafish juveniles in each group.

[0151]

[0152] Note: Data are expressed as mean ± standard deviation (n=3). Compared with the CON group, * P<0.05,** P<0.01 and *** P<0.001.

[0153] Currently, clinical treatments for psoriasis primarily involve topical medications such as retinoids, vitamin D3 analogs, and corticosteroids. These drugs mainly target the pathological processes of excessive epidermal keratinocyte proliferation and inflammatory cell infiltration during the development of psoriatic lesions. For example, retinoids and vitamin D3 analogs can inhibit excessive epidermal keratinocyte proliferation, while corticosteroids can suppress excessive inflammatory responses. However, these drugs all have varying degrees of side effects, such as skin irritation and contraindications for long-term use. In particular, long-term use of corticosteroids can cause skin atrophy and telangiectasia. Furthermore, research on topical medications targeting another pathological aspect of psoriatic lesion development—skin angiogenesis—is lacking.

[0154] This invention provides a composition of baicalin and echinacoside, natural active ingredients derived from medicinal plants. Studies have confirmed that these two natural plant active ingredients are essentially non-toxic to cells and have higher safety compared to glucocorticoids and retinoids, making them suitable for long-term use. Studies in Examples 3 and 4 revealed that baicalin can inhibit the release of inflammatory factors such as TNF-α through the TNF signaling pathway and inhibit macrophage inflammatory infiltration; this effect is superior to echinacoside to some extent. Echinacoside can inhibit the expression of key angiogenesis factors such as VEGF-A through the VEGF signaling pathway and inhibit angiogenesis by inhibiting the expression of another key angiogenesis factor, MMP-9, while also reducing oxidative damage; this effect is also superior to baicalin to some extent. Furthermore, the synergistic effect of the combination of baicalin and echinacoside is achieved through combination 2 (baicalin:echinacoside = 2:1), which shows a significantly better effect than single components of baicalin and echinacoside.

Claims

1. A composition of active substances from a traditional Chinese medicine with anti-psoriasis effects, characterized in that, The composition is: 2 parts by weight of baicalin and 1 part by weight of echinacoside.

2. A pharmaceutical preparation, characterized in that: A composition containing the active traditional Chinese medicine substances with anti-psoriasis effects as described in claim 1.

3. The pharmaceutical preparation according to claim 2, characterized in that: The preparation described is for external use.

4. The pharmaceutical preparation according to claim 3, characterized in that: The topical preparation is a cream.

5. The pharmaceutical preparation according to claim 4, characterized in that, The composition includes the following weight percentages of matrix excipients and active substances from traditional Chinese medicine. Oil phase: Stearic acid 8~12% Glyceryl monostearate 4-8% 3-4% white petrolatum Aqueous phase: Triethanolamine 4-6% Glycerol 4-8% Azone 1~3% 4-8% of the active ingredients in traditional Chinese medicine Ethylparaben 0.05~0.2% Ultrapure water balance The active ingredient composition of the traditional Chinese medicine is a mixture of baicalin and echinacoside.

6. The method for preparing the pharmaceutical formulation according to claim 4 or 5, characterized in that, Includes the following steps: (1) Heat 8-12% stearic acid, 4-8% glyceryl monostearate and 3-4% white petrolatum in a water bath to 78-82°C to melt them into a liquid to form an oil phase; (2) 4-6% triethanolamine, 4-8% glycerol, 1-3% azone, 4-8% active traditional Chinese medicine composition, 0.05-0.2% ethylparaben and an appropriate amount of ultrapure water are heated in a water bath to 78-82°C to completely dissolve them into a solution, forming an aqueous phase; (3) Add the oil phase to the aqueous phase, then add the remaining ultrapure water, emulsify, and the cream is obtained.

7. The use of the active ingredient composition of traditional Chinese medicine according to claim 1 in the preparation of an anti-psoriasis topical cream.

8. The use of the pharmaceutical preparation according to claim 2 in the preparation of an anti-psoriasis topical cream.

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