Pharmaceutical use of a white flower spatholobi extract

Abstract

The present invention relates to the technical field of natural drug chemistry, and specifically discloses an application of a petroleum ether extract of Eleutherococcus scabra in preparing an osteoclast differentiation inhibitor. The osteoclast differentiation inhibitor is used to prepare one or more drugs for preventing and / or treating osteoporosis, osteolysis, periprosthetic bone dissolution, tumor bone metastasis, inflammatory bone loss, and periodontitis inflammatory bone resorption diseases.

Description

Technical Field

[0001] The invention belongs to the technical field of biomedicine, and particularly relates to application of a petroleum ether extract of Eleutherococcus scabra in preparing an osteoclast differentiation inhibitor. Background Art

[0002] Bone metabolism mainly includes two processes: bone resorption by osteoclasts and bone formation by osteoblasts. When osteoclasts are overactivated, that is, bone resorption is greater than bone formation, the original bone metabolism balance is broken, and symptoms such as decreased bone density and bone mass will occur. Overactivation of osteoclasts is the main pathogenic mechanism of diseases such as periprosthetic bone dissolution, osteoporosis, and tumor bone metastasis.

[0003] Artificial joint replacement is an important method for the treatment of osteoarthritis in clinical practice. It can effectively relieve joint pain and has the effect of rebuilding joint function. However, periprosthetic bone dissolution leads to aseptic loosening of the prosthesis, which is the main complication after artificial joint replacement and the main reason for artificial joint revision. It is currently a hot topic and difficulty in orthopedics.

[0004] In osteoporosis, due to the high activity of osteoclast bone resorption in bone metabolism, osteoblast bone formation slows down, resulting in decreased bone mass and degradation of bone tissue microstructure (cancellous bone trabeculae become thinner, broken, and reduced in number; cortical bone becomes porous and thinner), resulting in increased bone fragility and increased risk of fracture. Therefore, inhibiting osteoclast-mediated bone resorption is an effective way to treat osteoporosis. However, bone resorption inhibitors currently used in clinical practice, such as bisphosphonates, have an increased risk of causing atypical femoral fractures. Therefore, it is of great significance and value to study new drugs that inhibit bone resorption from traditional Chinese medicine.

[0005] Elephantopus tomentosus L. is the dried whole herb of Elephantopus tomentosus L., a plant of the genus Elephantopus in the Asteraceae family. It is also known as ox tongue grass and is mainly distributed in tropical areas such as South China and Southwest China, such as Fujian and Guangdong. The whole herb of Elephantopus tomentosus is used as medicine. It is cool in nature, pungent and bitter in taste, and has the effects of clearing heat and detoxifying, anti-inflammatory, cooling blood, promoting dampness and reducing swelling. It is mainly used clinically to treat diseases such as tonsillitis, colds, hepatitis, and carbuncle. However, there is little research on this plant at present. Therefore, searching for the active ingredients from the traditional Chinese medicine Elephantopus tomentosus has broad application prospects and important application value for the development of drugs for the prevention and treatment of bone-related diseases such as prosthetic loosening and osteolysis, osteoporosis, and tumor bone metastasis. Summary of the invention

[0006] The present invention aims to provide a new pharmaceutical use of an extract of Eleutherodactylum scabra, namely, the use of an extract of Eleutherodactylum scabra petroleum ether in the preparation of an osteoclast differentiation inhibitor. The use of the extract of Eleutherodactylum scabra petroleum ether in the preparation of an osteoclast differentiation inhibitor is first discovered and has the advantages of significant effect and the like.

[0007] Furthermore, the preparation method of the petroleum ether extract of the white flower scutellaria comprises the following steps: (1) taking the traditional Chinese medicine white flower scutellaria, extracting it with an ethanol solution, concentrating the extract and removing the ethanol to obtain a total extract of the white flower scutellaria; (2) suspending the total extract of the white flower scutellaria in water, letting it stand, and then adding petroleum ether to extract to obtain the petroleum ether extract of the white flower scutellaria.

[0008] Furthermore, the ethanol solution is an ethanol aqueous solution with a volume fraction of 70-95%. Preferably, the ethanol solution is an ethanol aqueous solution with a volume fraction of 95%.

[0009] Furthermore, the ratio of the amount of Elaeagnus alba to the ethanol solution is 1 g: 8-20 mL. Preferably, the ratio of the amount of Elaeagnus alba to the ethanol solution is 1 g: 15 mL.

[0010] Furthermore, the extraction in step (1) refers to extraction by cold soaking method, and the extraction time is 3 to 10 days. Preferably, the extraction time is 5 days.

[0011] Furthermore, in step (2), the ratio of the total extract of Elaeagnus alba to water is 1 g: 15-30 mL. Preferably, in step (2), the ratio of the total extract of Elaeagnus alba to water is 1 g: 20 mL.

[0012] Furthermore, in step (2), the volume ratio of petroleum ether to water is 1 to 3:1, preferably 2:1.

[0013] Furthermore, the osteoclast differentiation inhibitor is used to prepare one or more drugs for preventing and / or treating osteoporosis, osteolysis, periprosthetic bone dissolution, tumor bone metastasis, inflammatory bone loss, and periodontitis inflammatory bone resorption diseases.

[0014] Advantages and beneficial effects of the present invention: The application of the petroleum ether extract of Eleutherococcus scabra in the preparation of osteoclast differentiation inhibitors is discovered for the first time, and its inhibitory effect on osteoclast differentiation is significantly better than that of the prior art. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 A schematic diagram showing the effects of Examples 1 to 5 and Comparative Examples 1 to 5 provided by the present invention on osteoclast differentiation of osteoclast precursor cells, i.e., macrophage cell line RAW264.7 cells;

[0016] Figure 2A schematic diagram of the cytotoxicity results of Example 1 and Comparative Examples 1 to 3 provided by the present invention on primary mouse macrophages BMMs cells;

[0017] Figure 3 A schematic diagram showing the effects of Example 1 and Comparative Examples 1 to 3 provided by the present invention on osteoclast differentiation of primary mouse macrophages BMMs cells;

[0018] Figure 4 A representative Micro-CT image of the inhibitory effect of Example 1 on osteolysis provided by the present invention;

[0019] Figure 5 This is a quantitative schematic diagram of the inhibitory effect of Example 1 on osteolysis provided by the present invention. DETAILED DESCRIPTION

[0020] The present invention is further explained below in conjunction with specific examples, but the examples do not limit the present invention in any form.

[0021] Example 1 Preparation of the petroleum ether extract of Euphorbia pulegata

[0022] The Chinese medicinal material of Herba Epimedii is extracted by cold soaking with 95% ethanol-water solution by volume fraction, and the extract is concentrated and the ethanol is removed to obtain the Herba Epimedii total extract; the Herba Epimedii total extract is suspended in water, and petroleum ether is added to extract the Herba Epimedii petroleum ether extract after standing. The cold soaking extraction time is 5 days, the dosage ratio of Herba Epimedii to ethanol is 1 g:15 mL, the dosage ratio of Herba Epimedii total extract to water is 1 g:20 mL, and the volume dosage ratio of petroleum ether to water is 2:1.

[0023] Example 2 Preparation of the petroleum ether extract of Euphorbia pulegosa

[0024] The Chinese medicinal material of Herba Epimedii was extracted by cold soaking with 95% ethanol-water solution by volume fraction, and the extract was concentrated and the ethanol was removed to obtain the Herba Epimedii total extract; the Herba Epimedii total extract was suspended in water, and petroleum ether was added to extract the Herba Epimedii petroleum ether extract after standing. The cold soaking extraction time was 3 days, the dosage ratio of Herba Epimedii to ethanol was 1 g: 8 mL, the dosage ratio of Herba Epimedii total extract to water was 1 g: 15 mL, and the volume dosage ratio of petroleum ether to water was 1: 1.

[0025] Example 3 Preparation of the petroleum ether extract of Euphorbia pulegata

[0026] The Chinese medicinal material of Herba Epimedii was extracted by cold soaking with 70% ethanol-water solution by volume fraction, and the extract was concentrated and the ethanol was removed to obtain the Herba Epimedii total extract; the Herba Epimedii total extract was suspended in water, and petroleum ether was added to extract the Herba Epimedii petroleum ether extract after standing. The cold soaking extraction time was 3 days, the dosage ratio of Herba Epimedii to ethanol was 1 g: 8 mL, the dosage ratio of Herba Epimedii total extract to water was 1 g: 15 mL, and the volume dosage ratio of petroleum ether to water was 1: 1.

[0027] Example 4 Preparation of the petroleum ether extract of Euphorbia pulegata

[0028] The Chinese medicinal material of Herba Epimedii is extracted by cold soaking with an ethanol-water solution having a volume fraction of 80%, and the extract is concentrated and the ethanol is removed to obtain a Herba Epimedii total extract; the Herba Epimedii total extract is suspended in water, and petroleum ether is added to extract the Herba Epimedii petroleum ether extract after standing. The cold soaking extraction time is 10 days, the dosage ratio of Herba Epimedii to ethanol is 1 g: 20 mL, the dosage ratio of Herba Epimedii total extract to water is 1 g: 30 mL, and the volume dosage ratio of petroleum ether to water is 3: 1.

[0029] Example 5 Preparation of petroleum ether extract of Euphorbia pulegata

[0030] The Chinese medicinal material of Herba Epimedii was extracted by cold soaking with 95% ethanol-water solution by volume fraction, and the extract was concentrated and the ethanol was removed to obtain the Herba Epimedii total extract; the Herba Epimedii total extract was suspended in water, and petroleum ether was added to extract the Herba Epimedii petroleum ether extract after standing. The cold soaking extraction time was 10 days, the dosage ratio of Herba Epimedii to ethanol was 1 g: 20 mL, the dosage ratio of Herba Epimedii total extract to water was 1 g: 30 mL, and the volume dosage ratio of petroleum ether to water was 3: 1.

[0031] Comparative Example 1 Preparation of dichloromethane extract of Euphorbia pulegata

[0032] The Chinese medicinal material of the herb of white flower dandelion is extracted by cold soaking with an ethanol-water solution having a volume fraction of 95%, and the extract is concentrated and the ethanol is removed to obtain the total extract of the herb of white flower dandelion; the total extract of the herb of white flower dandelion is suspended in water, and after standing, dichloromethane is added to extract the dichloromethane extract of the herb of white flower dandelion. The cold soaking extraction time is 5 days, the dosage ratio of the herb of white flower dandelion to ethanol is 1g:15mL, the dosage ratio of the total extract of the herb of white flower dandelion to water is 1g:20mL, and the volume dosage ratio of dichloromethane to water is 2:1.

[0033] Comparative Example 2 Preparation of Hexane Extract of Euphorbia pulegata

[0034] The Chinese medicinal material of Herba Epimedii is extracted by cold soaking with 95% ethanol-water solution by volume fraction, and the extract is concentrated and the ethanol is removed to obtain the Herba Epimedii total extract; the Herba Epimedii total extract is suspended in water, and after standing, n-hexane is added to extract the Herba Epimedii n-hexane extract. The cold soaking extraction time is 5 days, the dosage ratio of Herba Epimedii to ethanol is 1 g: 15 mL, the dosage ratio of the Herba Epimedii total extract to water is 1 g: 20 mL, and the volume dosage ratio of n-hexane to water is 2: 1.

[0035] Comparative Example 3 Preparation of Cyclohexane Extract of Euphorbia pulegata

[0036] The Chinese medicinal material of the herb of white flower dandelion is extracted by cold soaking with an ethanol-water solution having a volume fraction of 95%, and the extract is concentrated and the ethanol is removed to obtain the total extract of the herb of white flower dandelion; the total extract of the herb of white flower dandelion is suspended in water, and after standing, cyclohexane is added to extract the cyclohexane extract of the herb of white flower dandelion. The cold soaking extraction time is 5 days, the dosage ratio of the herb of white flower dandelion to ethanol is 1g:15mL, the dosage ratio of the total extract of the herb of white flower dandelion to water is 1g:20mL, and the volume dosage ratio of cyclohexane to water is 2:1.

[0037] Comparative Example 4 Preparation of Ethyl Acetate Extract of Euphorbia pulegata

[0038] The Chinese medicinal material of white flower dandelion is extracted by cold soaking method with 95% ethanol water solution by volume fraction, and the extract is concentrated and then ethanol is removed to obtain the total extract of white flower dandelion; the total extract of white flower dandelion is suspended in water, and petroleum ether is added to extract the petroleum ether extract of white flower dandelion after standing; and ethyl acetate solution is continuously added to the remaining part to extract the ethyl acetate extract of white flower dandelion. The cold soaking extraction time is 5 days, the dosage ratio of white flower dandelion to ethanol is 1g:15mL, the dosage ratio of white flower dandelion total extract to water is 1g:20mL, the volume dosage ratio of petroleum ether to water is 2:1, and the volume dosage ratio of ethyl acetate solution to water is 2:1.

[0039] Comparative Example 5 Preparation of the n-butanol extract of Eleutherodactylum scabra.

[0040] The Chinese medicinal material of white flower dandelion is extracted by cold soaking method with 95% ethanol aqueous solution by volume fraction, and the extract is concentrated and then ethanol is removed to obtain the total extract of white flower dandelion; the total extract of white flower dandelion is suspended in water, and petroleum ether is added to extract the petroleum ether extract of white flower dandelion after standing; ethyl acetate solution is continuously added to the remaining part to extract the ethyl acetate extract of white flower dandelion; finally, n-butanol is continuously added to the remaining part after ethyl acetate extraction to obtain the n-butanol extract of white flower dandelion. Among them, the cold soaking extraction time is 5 days, the dosage ratio of white flower dandelion to ethanol is 1g:15mL, the dosage ratio of white flower dandelion total extract to water is 1g:20mL, the volume dosage ratio of petroleum ether to water is 2:1, the volume dosage ratio of ethyl acetate solution to water is 2:1, and the volume dosage ratio of n-butanol solution to water is 2:1.

[0041] Experimental part: The extracts of Euphorbia pulegium prepared in Examples 1 to 5 and Comparative Examples 1 to 5 were dissolved in DMSO to prepare a 1 mg / mL test solution; used to test the osteoclast differentiation-inhibiting activities of the extracts prepared in Examples 1 to 5 and Comparative Examples 1 to 5.

[0042] Experimental Example 1 Experimental study on the effect of the extract of Euphorbia pulex on the differentiation and maturation of osteoclast precursor cells, i.e. macrophage cell line RAW264.7

[0043] Take RAW264.7 cells in the logarithmic growth phase, plant RAW264.7 cells at a density of 3000 cells / well in a 96-well plate, and culture them in an α-MEM medium containing 10% FBS (fetal bovine serum) and 1% antibiotics; after 6 hours of culture, replace the medium with osteoclast induction factor M-CSF (20 ng / mL), RANKL (5ng / mL) (USA, R&D System) and different concentrations of white flowered dandelion extracts prepared in Examples 1 to 5 and Comparative Examples 1 to 5 (1μg / mL, 10μg / mL, 100μg / mL, 1000μg / mL), set 3 replicates in each group, culture for 4 days, and replace new culture medium and drugs every 2 days. After a total of 4 days of culture, remove the culture medium containing the compound, aspirate the culture medium, rinse once with cold PBS, fix with 4% paraformaldehyde solution, and finally use TRAP, i.e., tartrate-resistant acid phosphatase, for staining. The nuclei of the induced mature multinucleated osteoclasts can be stained dark by TRAP staining reagent, so the cells with more than 3 nuclei produced by differentiation were counted and the area was quantified under a microscope.

[0044] Test results such as Figure 1As shown. The results show that the petroleum ether extract of Example 1 showed an inhibitory effect on the number of multinuclear osteoclasts at two concentrations of 100ng / mL and 1000ng / mL, and the difference with the control group was statistically significant (***p<0.001); the petroleum ether extracts of Examples 2 to 5 showed a certain inhibitory effect on the number of multinuclear osteoclasts at a concentration of 1000ng / mL, and the difference with the control group was statistically significant (*p<0.05, **p<0.01, ***p<0.001); the dichloromethane extract of Comparative Example 1 and the n-hexane extract of Comparative Example 2 showed a certain inhibitory effect on the number of multinuclear osteoclasts at a concentration of 1000ng / mL, and the difference with the control group was statistically significant (*p<0.05); the other experimental groups did not show significant inhibitory effects. The comparison results show that the inhibitory effect of the petroleum ether extract of Example 1 on the number of osteoclasts is significantly better than that of the dichloromethane extract of Comparative Example 1 and the n-hexane extract of Comparative Example 2.

[0045] Experimental Example 2 Example 1 Cytotoxicity of the petroleum ether extract of Eleutherococcus scabra on primary mouse bone marrow mononuclear cells (BMMs)

[0046] Primary mononuclear cells were extracted from the bone marrow of 4-6 week-old C57BL / 6J mice and incubated at 37°C with 5% CO 2 After culturing overnight in the incubator, the supernatant was collected, centrifuged, and the cells were resuspended. 6 The cells were seeded at a density of 1 μg / mL in a 96-well plate and cultured in an α-MEM medium containing 10% FBS (fetal bovine serum), 1% antibiotics and M-CSF (20 ng / ml); after culturing for 24 hours, different concentrations of the petroleum ether extract of the white flowered dandelion prepared in Example 1 (1 μg / mL, 10 μg / mL, 100 μg / mL, 1000 μg / mL) were added, and 3 replicate wells were set for each group, and the culture was carried out for 4 days. New culture medium and drugs were replaced every 2 days. After a total of 4 days of culture, the culture medium containing the compound was removed, and an α-MEM culture medium containing 10 μL of CCK-8 reagent was added to each well, and the cells were incubated at 5% CO. 2 , incubate in a 37 °C incubator for 2 h, shake at room temperature for 10 s, and measure the absorbance (OD) of each well at 450 nm using a microplate reader.

[0047] Test results such as Figure 2 The results showed that compared with the control group, the petroleum ether extract of Eleutherococcus aviculare in Example 1 had no obvious cytotoxicity (p>0.05).

[0048] Experimental Example 3 Effect of the extract of Eleutherodactylum leucoderma on differentiation and maturation of primary bone marrow mononuclear cells (BMMs) of mice

[0049] Primary mononuclear cells were extracted from the bone marrow of 4-6 week-old C57BL / 6J mice and incubated at 37°C with 5% CO 2 After culturing overnight in the incubator, the supernatant was collected, centrifuged, and the cells were resuspended. 6 The cells were seeded at a density of 100 / mL in a 96-well plate. After culturing for 1 day in α-MEM complete medium containing 20 ng / mL M-CSF, the medium was replaced with α-MEM complete medium containing osteoclast induction factor M-CSF (20 ng / mL), RANKL (15 ng / mL) and different concentrations of white flower dandelion extract (1μg / mL, 10μg / mL, 100μg / mL, 1000μg / mL). Three replicate wells were set up in each group, and new medium and drugs were replaced every 2 days. After 5 days of administration, the medium containing the compound was removed, rinsed with cold PBS, fixed with 4% paraformaldehyde solution, and TRAP staining was performed with the TRAP detection kit. The nuclei of mature multinucleated osteoclasts induced can be dyed wine red by the staining solution. Finally, the cells with more than 3 nuclei produced by fusion were counted and the area was quantified under a visible light inverted microscope.

[0050] Test results such as Figure 3 The results show that Figure 3 In (A and C), the blank control group was used as the control, and the number of osteoclasts in the control group increased significantly. Among them, the petroleum ether extract of Example 1 and the n-hexane extract of Comparative Example 2 showed a significant inhibitory effect on the number of multinuclear osteoclasts at a concentration of 1000 ng / mL (**p<0.01, ***p<0.001), among which the petroleum ether extract of Example 1 had a better inhibitory effect on the number of osteoclasts than the n-hexane extract of Comparative Example 2, while other test groups such as the dichloromethane extract of Comparative Example 1 and the cyclohexane extract of Comparative Example 3 had no significant inhibitory effect on the number of osteoclasts. Figure 3 In (B and C), the blank control group was used as the control. The osteoclast area of ​​the control group increased significantly. The petroleum ether extract of Example 1 and the n-hexane extract of Comparative Example 2 showed a significant inhibitory effect on the area of ​​multinuclear osteoclast formation at a concentration of 1000 ng / mL (**p<0.01, ***p<0.001). The inhibitory effect of the petroleum ether extract of Example 1 on the osteoclast area was significantly better than that of the n-hexane extract of Comparative Example 2, while the other groups such as the dichloromethane extract of Comparative Example 1 and the cyclohexane extract of Comparative Example 3 had no significant inhibitory effect on the osteoclast area.

[0051] The healthy male C57BL / 6J mice used in the following experimental examples had an animal license number of SYXK (Guangdong) 2022-0174 and were purchased from Guangdong Weitong Lihua Experimental Animal Technology Co., Ltd.

[0052] Experimental Example 4 Example 1 Experiment on the inhibitory effect of petroleum ether extract of Eleutherococcus scabra on skull osteolysis induced by titanium particles

[0053] We further selected the petroleum ether group that inhibited osteoclast differentiation activity better than other groups in the in vitro cell experiment to carry out the in vivo cranial bone dissolution inhibitory activity evaluation test. 8-week-old male C57BL / 6J mice, weighing 25±2g, were selected and raised in an SPF animal room with free access to food and water. After one week of adaptive feeding, the mice were randomly divided into 4 groups: sham operation group, osteolysis model group, low-dose group of petroleum ether extract in Example 1, and high-dose group of petroleum ether extract in Example 1, with 8 mice in each group, for a total of 32 mice.

[0054] After the mice were anesthetized with isoflurane, the hair on the top of the skull was shaved with a shaver, and the skin on the top of the skull was disinfected with 75% ethanol. Then, a 1.5 cm long incision was made along the midline of the skull with a scalpel. 30 mg of Ti particles (30 μL) were evenly dropped on the surface of the skull, and the surgical incision was sutured to establish a mouse skull osteolysis model. The same volume of sterile PBS was dropped on the surface of the skull in the sham operation group. The drug was intraperitoneally injected on the second day after surgery. The sham operation group and the osteolysis model group were intraperitoneally injected with an equal amount of drug solvent, once a day, for 14 consecutive days. After 14 days, the experimental animals were killed, and the skull specimens were collected. After being fixed with 4% paraformaldehyde for 48 h, the Ti particles on the surface of the skull were carefully removed. A high-resolution Micro-CT instrument (Skyscan 1176; Bruker, Antwerp, Belgium) was used with a scanning resolution of 10.4 μm, an X-ray voltage of 55 kV, and a current of 145 μA. The circular area centered on the median suture of the skull was used as the region of interest (ROI). Bone mineral density (BMD), bone volume fraction (BV / TV), bone surface area to bone volume ratio (BS / BV) and trabecular thickness (Tb. Th) were calculated using CTAn software; 3D reconstructed images were constructed using CTvox.

[0055] The results are as follows Figure 4 and Figure 5 As shown. The results showed that compared with the sham operation group, the osteolysis model group caused a large amount of bone loss in the skull of mice, BMD, BV / TV and Th.Tb were significantly reduced, BS / BV was significantly increased, and the difference was statistically significant (***p<0.001). After the high-dose group of petroleum ether extract in Example 1 was administered, the bone loss of the skull of mice was significantly improved. Compared with the osteolysis model group, BMD, BV / TV and Th.Tb were significantly increased, BS / BV was significantly reduced, and the difference was statistically significant (#p<0.05, ##p<0.01), and no significant improvement was shown in other groups.

[0056] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention rather than to limit the protection scope of the present invention. For ordinary technicians in this field, other different forms of changes or modifications can be made based on the above descriptions and ideas. It is not necessary and impossible to list all the implementation methods here. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the protection scope of the claims of the present invention.

Claims

1. Use of the petroleum ether extract of Euphorbia pulegiae in the preparation of one or more drugs for preventing and / or treating osteoporosis, osteolysis, and tumor bone metastasis, characterized in that: The preparation method of the petroleum ether extract of Elaeagnus alba comprises the following steps: (1) extracting Elaeagnus alba from a traditional Chinese medicine with an ethanol solution, concentrating the extract and removing the ethanol to obtain a total extract of Elaeagnus alba, wherein the ethanol solution is an ethanol aqueous solution with a volume fraction of 70 to 95%; (2) suspending the total extract of Elaeagnus alba with water, allowing it to stand, and then adding petroleum ether to extract the petroleum ether extract of Elaeagnus alba.

2. The use according to claim 1, characterized in that: The extraction in step (1) refers to extraction by cold soaking method, the extraction time is 3 to 10 days, and the dosage ratio of white flowered dandelion to ethanol solution is 1g:8 to 20mL.

3. The use according to claim 2, characterized in that: The extraction time in step (1) is 5 days, the ethanol solution is an ethanol aqueous solution with a volume fraction of 95%, and the dosage ratio of the white flower dandelion to the ethanol solution is 1g:15mL.

4. The use according to any one of claims 1 to 3, characterized in that: In step (2), the ratio of the total extract of Euphorbia alba to water is 1 g:15-30 mL, and the ratio of the volume of petroleum ether to the volume of water is 1-3:

1.

5. The use according to claim 4, characterized in that: In step (2), the ratio of the total extract of Euphorbia alba to water is 1 g:20 mL, and the ratio of the volume of petroleum ether to the volume of water is 2:1.

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