Use of salvia miltiorrhiza extract in preparation of aromatase inhibitors

Abstract

The application relates to application of a salvia miltiorrhiza extract in preparation of an aromatase inhibitor. Compared with the existing synthetic aromatase inhibitor, the salvia miltiorrhiza extract is natural, has high safety, small side effect, and has good treatment effect and important clinical application prospect in treatment and prevention of estrogen-dependent diseases such as breast cancer, breast hyperplasia, endometrial hyperplasia and the like.

Description

Technical Field

[0001] This invention relates to the field of pharmaceuticals, and more specifically to the application of tanshinone extract in the preparation of aromatase inhibitors. Background Technology

[0002] Aromatase, a member of the cytochrome P450 family, primarily functions to catalyze the conversion of androgens to estrogens. This enzyme is widely distributed in the central nervous system, ovaries, adipose tissue, and skeletal muscle. In women (especially postmenopausal women), estrogen is produced in peripheral fat, muscle, and breast tissue, particularly in breast tissue. High estrogen levels can induce tumor proliferation. Inhibiting aromatase can suppress estrogen production; therefore, aromatase has become a key target for breast cancer treatment.

[0003] Aromatase inhibitors are a class of drugs that block the conversion of androgens into estrogens by inhibiting aromatase activity. Aromatase inhibitors can effectively reduce estrogen levels in tissues and inhibit the growth of breast cancer cells. Currently, commonly used aromatase inhibitors in clinical practice include letrozole, anastrozole, and exemestane. However, existing aromatase inhibitors are prone to side effects such as musculoskeletal damage, endocrine disorders, and metabolic abnormalities with long-term use, thus limiting their widespread use. Therefore, there is still an urgent need to develop an aromatase inhibitor with high safety, low side effects, and suitability for long-term use.

[0004] Danshen (Salvia miltiorrhiza) is a plant belonging to the genus *Salvia* in the family Lamiaceae, and it is widely used in Traditional Chinese Medicine. Its main components include various natural products, and its main effects include promoting blood circulation, reducing inflammation, and acting as an antioxidant. It is primarily used to treat cardiovascular diseases. However, there are currently no studies or reports on the use of Danshen and its extracts as aromatase inhibitors for the treatment or prevention of estrogen-dependent diseases. Summary of the Invention

[0005] To solve the above-mentioned technical problems, the present invention includes the following aspects: A first aspect of the present invention provides the use of tanshinone extract in the preparation of aromatase inhibitors.

[0006] Preferably, the present invention provides the application of tanshinone extract as the sole active ingredient in the preparation of aromatase inhibitors.

[0007] Preferably, the preparation method of the tanshinone extract is as follows: (1) Weigh out the prescribed amount of Danshen medicinal material, pulverize it and sieve it to obtain medicinal material powder; (2) Add extraction solvent to the medicinal powder obtained in step (1) and extract by reflux to obtain Danshen extract; (3) The extract obtained in step (2) is concentrated and dried to obtain the extract of tanshinone.

[0008] Preferably, the danshen in step (1) is the root or underground part of danshen.

[0009] Preferably, in step (1), the pulverized Salvia miltiorrhiza is passed through a 40-60 mesh sieve.

[0010] Preferably, in step (2), the ratio of medicinal powder to extraction solvent is 1:5-1:20 (kg / L). More preferably, in step (2), the ratio of medicinal powder to extraction solvent is 1:10 (kg / L).

[0011] Preferably, the extraction solvent in step (2) is methanol or a 95% (v / v) ethanol solution.

[0012] Preferably, the reflux extraction in step (2) is performed by heating and refluxing at 70-90℃ 2-3 times, with each extraction time being 80-120 min. More preferably, the reflux extraction in step (2) is performed by heating and refluxing at 80℃ 3 times, with each extraction time being 100 min.

[0013] Preferably, the concentration in step (3) is carried out by rotary evaporation at -0.08 to -0.09 MPa and 55 to 65°C.

[0014] Preferably, the drying in step (3) is to dry the concentrated tanshinone extract at 60°C to a tanshinone extract paste with a relative density of 1.10-1.25.

[0015] Preferably, the preparation method of the tanshinone extract is as follows: (1) Weigh out the prescribed amount of Salvia miltiorrhiza root, crush it and sieve it to obtain medicinal powder; (2) Add 95% (v / v) ethanol solution to the medicinal powder obtained in step (1) at a material-to-liquid ratio of 1:10 (kg / L), and heat and reflux at 80°C for 3 times, with each extraction time being 100 min, to obtain Danshen extract; (3) The extract obtained in step (2) is concentrated and dried to obtain the extract of tanshinone.

[0016] Preferably, the tanshinone extract is total phenolic acid from tanshinone.

[0017] Preferably, the present invention provides the application of total phenolic acids from tanshinone as the sole active ingredient in the preparation of aromatase inhibitors.

[0018] Preferably, the method for preparing the total phenolic acids of Tanshinone is as follows: (1) Weigh out the prescribed amount of Danshen medicinal material, pulverize it and sieve it to obtain medicinal material powder; (2) Add water to the herbal powder and heat to extract. Adjust the pH of the extract to 2 with hydrochloric acid and filter. (3) Load the filtrate onto a polyamide column, wash with deionized water, then elute with sodium bicarbonate aqueous solution, and collect the eluent; (4) Adjust the pH of the eluent to 2 with hydrochloric acid, then load the eluent onto the macroporous adsorption resin, rinse with deionized water until neutral, and continue eluting with 95% ethanol. Collect the band when it comes down. (5) Concentrate the collected liquid under reduced pressure until it is completely dry, dissolve it in an appropriate amount of water, and then filter it through a microporous membrane to obtain the total phenolic acid extract of tanshinone. (6) Adjust the pH of the extract to 6.0 with sodium hydroxide solution, freeze dry, and obtain total phenolic acid lyophilized powder of tanshinone.

[0019] Preferably, the water extraction step in step (2) is as follows: water is added and heated for extraction 3 times. For the first extraction, 5.5 times the weight of the medicinal material is added to water and heated for 1 hour. For the second and third extractions, 3 times the weight of the medicinal material is added to water and heated for 0.5 hours each time.

[0020] Preferably, in step (3), the sample is first washed with 5 column volumes of deionized water, and then eluted with 5 column volumes of 0.1% sodium bicarbonate aqueous solution.

[0021] Preferably, the macroporous adsorption resin in step (4) is D. 101 Macroporous adsorption resin.

[0022] Preferably, the microporous filter membrane in step (5) is a 0.3 μm mixed cellulose microporous filter membrane.

[0023] A second aspect of the present invention provides the use of a pharmaceutical composition in the preparation of an aromatase inhibitor, said pharmaceutical composition comprising the tanshinone extract described in the first aspect.

[0024] Preferably, the dosage form of the pharmaceutical composition is selected from oral liquid, tablet, granule, capsule or injection.

[0025] Preferably, the pharmaceutical composition further comprises other aromatase inhibitors.

[0026] Preferably, the other aromatase inhibitor is selected from one or more of ammonia glutide, formestan, letrozole, anastrozole, and exemestane.

[0027] A third aspect of the present invention provides the use of the tanshinone extract described in the first aspect or the pharmaceutical composition described in the second aspect in the preparation of a medicament for the prevention or treatment of estrogen-dependent diseases.

[0028] Preferably, the present invention provides the use of tanshinone extract as the sole active ingredient in the preparation of a medicament for the prevention or treatment of estrogen-dependent diseases.

[0029] Preferably, the estrogen-dependent disease is selected from breast cancer, breast hyperplasia, endometrial cancer, endometrial hyperplasia, endometriosis, uterine leiomyomas, or gynecomastia.

[0030] More preferably, the breast cancer is estrogen receptor-positive breast cancer in postmenopausal women.

[0031] Preferably, the tanshinone extract is used to treat or prevent estrogen-dependent diseases by inhibiting aromatase activity.

[0032] The technical effects of this invention are as follows: This invention unexpectedly discovered that tanshinone extract and total phenolic acids from tanshinone can effectively inhibit aromatase activity and reduce estrogen levels in the body, thus they can be used as aromatase inhibitors. Compared with existing synthetic aromatase inhibitors, the tanshinone extract and total phenolic acids from tanshinone of this invention are naturally derived, have high safety, and few side effects, showing good therapeutic effects and significant clinical application prospects in the treatment and prevention of estrogen-dependent diseases such as breast cancer, breast hyperplasia, and endometrial hyperplasia. Attached Figure Description

[0033] Figure 1 This is the result of an experiment on the inhibition rate of aromatase by tanshinone extract; Figure 2 This is the result of an assay to test the inhibition rate of ammoniacal enzymes by aminoglutethimide; Figure 3 The results are from the test on the inhibition rate of total phenolic acids from tanshinone against aromatase. Figure 4 The results show the effect of gavage administration of Danshen extract on serum estradiol levels in ovariectomized mice. Asterisks indicate significant correlations: *P<0.05, **P<0.01, ***P<0.001. Figure 5 The results are from an experiment on the effect of tail vein injection of total phenolic acids from tanshinone on serum estradiol levels in ovariectomized mice. Figure 6 This is the result of an experiment on the effect of oral administration of total phenolic acids from tanshinone on the serum estradiol level in ovariectomized mice. Detailed Implementation

[0034] Unless otherwise specified, the methods involved in the different embodiments of the present invention described below are conventional methods, and the reagents used are conventional commercially available reagents or reagents prepared according to conventional methods, unless otherwise specified. Furthermore, the technical features involved in the different embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0035] Example 1: Preparation of Salvia miltiorrhiza extract 100g of dried Salvia miltiorrhiza root was weighed, impurities were removed, and the powder was pulverized and passed through a 40-60 mesh sieve to obtain medicinal powder. The powder was placed in a container, and 95% (v / v) ethanol solution was added at a material-to-liquid ratio of 1:10 (kg / L). The mixture was extracted three times by reflux at 80℃, with each extraction lasting 100 min. After extraction, the three extracts were combined, filtered while hot, and the filtrate was collected to obtain Salvia miltiorrhiza extract. The extract was then concentrated in a rotary evaporator at -0.08 to -0.09 MPa and 55 to 65℃. Concentration was stopped when the relative density of the extract reached 1.10-1.25 at 60℃. The concentrated product was dried to obtain 13.5g of brownish-yellow powdered Salvia miltiorrhiza extract, which was stored at -20℃ for later use.

[0036] Example 2: Preparation of total phenolic acids from Tanshinone 5 kg of Salvia miltiorrhiza was pulverized into coarse powder and extracted three times with water and heat. For the first extraction, 5.5 times the weight of the herb was added to water, and the mixture was heated for 1 hour. For the second and third extractions, 3 times the weight of the herb was added to water, and the mixture was heated for 0.5 hours each time. The extract was adjusted to pH 2 with hydrochloric acid and filtered. The filtrate was loaded onto a polyamide column, washed with 5 column volumes of deionized water, and then eluted with 5 column volumes of 0.1% sodium bicarbonate aqueous solution. The eluent was collected. The pH was adjusted to 2 with hydrochloric acid, and then the eluent was loaded onto a D... 101 The macroporous adsorption resin was rinsed with deionized water until neutral, and then eluted with 95% ethanol until a colored band appeared, which was then collected. The collected solution was concentrated under reduced pressure until completely dry, dissolved in an appropriate amount of water, and filtered through a 0.3 μm mixed cellulose microporous membrane to obtain the total phenolic acid extract of Tanshinone. The pH was adjusted to 6.0 with sodium hydroxide solution, and the solution was freeze-dried to obtain the lyophilized powder of total phenolic acids of Tanshinone.

[0037] Experimental Example 1: Determination of the inhibitory activity of tanshinone extract on aromatase 1. Test Methods In this experiment, aminoglutethimide was used as a positive control for aromatase inhibition activity. The in vitro aromatase inhibition activity of *Salvia miltiorrhiza* extract was determined using a 96-well cell plate. The experiment consisted of a blank control group, a positive control group, and a test sample group, with three parallel experimental groups in each group. The blank control group received 5 μL of DMSO, while the positive control group received 5 μL of aminoglutethimide solutions prepared with DMSO at concentrations of 0.5%, 1.0%, and 2.0%. The concentrations of the danshen extract prepared in Example 1 were 0.1, 0.5, 1.0, 2.0, 5.0, 10.0, 20.0, 50.0, and 75.0 μg / mL, respectively. 5 μL of the danshen extract solution prepared in DMSO was added to each sample group. The NADPH solution was preheated in a water bath for 10 min, and then 45 μL of NADPH solution was added to each well, followed by shaking and reaction in an incubator for 10 min. Simultaneously, the BSA solution containing DBF was preheated in a water bath. After 5 minutes, 2.5 μL of aromatase solution was added to the BSA solution to prepare the enzyme-substrate E / S system. After 7 minutes, the E / S system solution was heated in a water bath for 3 minutes. Then, 38 μL of NaOH solution was added to each well of the experimental group, followed by 50 μL of the E / S system solution. The 96-well cell plate was then placed in an incubator and shaken for 30 minutes. Then, 38 μL of NaOH solution was added to each well of the experimental group again, and the plate was placed in an incubator and shaken for 2 hours. Finally, the fluorescence intensity was measured at an excitation wavelength of 485 nm and a measurement wavelength of 528 nm using a microplate reader. The inhibition rate of aromatase by different concentrations of Tanshinone extract and aminoglutethimide was calculated according to the following formula: Aromatase inhibition rate = (1 - B / A) * 100% Where A represents the net fluorescence value of the blank group, and B represents the net fluorescence value of the positive control group or the sample group to be tested.

[0038] 2. Test Results like Figure 1 As shown, within the range of 0.1 to 75 μg / mL, the tanshinone extract of the present invention exhibits considerable inhibitory activity against aromatase, and the inhibition rate increases with increasing tanshinone extract concentration. Figure 2 As shown, aminoglutethimide exhibits considerable inhibitory activity against aromatase in the range of 0.1 to 80 μM, and the inhibition rate increases with increasing aminoglutethimide concentration.

[0039] The above data were statistically analyzed using GraphPad Prism 8.0 software, and the IC was obtained through nonlinear fitting. 50 The values ​​are expressed as mean ± standard deviation (SD). The IC50 of the tanshinone extract in this experimental case was calculated. 50 The IC50 of aminoglutethimide was 2.13 ± 0.15 μg / mL. 50 It is 10.49±0.67μM.

[0040] Experimental Example 2: Determination of the inhibitory activity of total phenolic acids from Tanshinone against aromatase. 1. Test Methods The inhibitory activity of total phenolic acids from Tanshinone against aromatase was determined according to the method in Example 1. The experiment was divided into a blank group and a test sample group, with three parallel experimental groups in each group. 5 μL of DMSO was added to the blank group, and 5 μL of a solution of total phenolic acids from Tanshinone prepared in Example 2, prepared in DMSO, was added to the test sample group. The concentrations were 0.1, 0.5, 1.0, 2.0, 4.0, 5.0, 7.5, 10.0, and 15.0 μg / mL, respectively. The NADPH solution was preheated in a water bath for 10 min, and then 45 μL of NADPH solution was added to each well, followed by shaking in an incubator for 10 min. Simultaneously, the BSA solution containing DBF was preheated in a water bath. After 5 minutes, 2.5 μL of aromatase solution was added to the BSA solution to prepare the enzyme-substrate E / S system. After 7 minutes, the E / S system solution was heated in a water bath for 3 minutes. Then, 38 μL of NaOH solution was added to each well of the experimental group, followed by 50 μL of the E / S system solution. The 96-well cell plate was then placed in an incubator and shaken for 30 minutes. Then, 38 μL of NaOH solution was added to each well of the experimental group again, and the plate was incubated again with shaking for 2 hours. Finally, the fluorescence intensity was measured at an excitation wavelength of 485 nm and a measurement wavelength of 528 nm using a microplate reader. The inhibition rate of aromatase by different concentrations of total phenolic acids from Tanshinone was calculated using the following formula: Aromatase inhibition rate = (1 - B / A) * 100% Where A represents the net fluorescence value of the blank group and B represents the net fluorescence value of the test sample group.

[0041] 2. Test Results like Figure 3 As shown, within the range of 0.1 to 15 μg / mL, the total phenolic acids from tanshinone of the present invention exhibit considerable inhibitory activity against aromatase, and the inhibition rate increases with increasing concentration of total phenolic acids from tanshinone. The IC50 of total phenolic acids from tanshinone in this experimental example was calculated. 50 It was 1.81 ± 0.56 μg / mL.

[0042] Experimental Example 3: Effect of Tanshinone Extract on Serum Estradiol Levels in Ovariectomized Mice 1. Test Methods Female BALB / c mice (20±5g, purchased from Spiford (Beijing) Biotechnology Co., Ltd.) were randomly divided into five groups: a blank control group, a model group, a positive control group, a low-dose tanshinone extract group, and a high-dose tanshinone extract group, with eight mice in each group. The tanshinone extract from Example 1 was added to a 0.5% CMC-Na solution as a solvent, and the extract was dispersed in the solvent using vortexing and ultrasonic dissolution methods. A 10mg / ml tanshinone extract suspension was prepared for the low-dose group, and a 30mg / ml tanshinone extract suspension was prepared for the high-dose group. Except for the blank control group, which underwent sham surgery, all mice in the other experimental groups underwent bilateral ovariectomy. Mice recovered for one week post-surgery to eliminate residual estrogen. After the recovery period, except for the blank control group, mice in all other experimental groups were subcutaneously injected with androstenedione at a dose of 0.5 mg / kg / day daily. Simultaneously, mice in each experimental group were administered the following medications via gavage for 21 consecutive days: the positive control group received letrozole at a dose of 1 mg / kg / day; the low-dose tanshinone extract group received a 10 mg / ml tanshinone extract solution at a dose of 100 mg / kg / day; the high-dose tanshinone extract group received a 30 mg / ml tanshinone extract solution at a dose of 300 mg / kg / day; and the blank control group and model group received an equal volume of 0.5% CMC-Na solution. Twenty-four hours after the last administration, blood was collected from the eyes of mice in each experimental group. Serum was separated by centrifugation, and the estradiol content in the serum of each group was detected using enzyme-linked immunosorbent assay (ELISA, purchased from Beijing Mairuida Technology Co., Ltd.).

[0043] 2. Test Results like Figure 4 As shown, compared with the blank control group, the serum estradiol level in the model group mice was significantly increased after injection of androstenedione (P<0.01), indicating that exogenous androgens were successfully converted into estrogens by aromatase in vivo. Compared with the model group, the serum estradiol levels in the positive control group, the high-dose tanshinone extract group, and the low-dose tanshinone extract group were all significantly decreased (P<0.05 or P<0.01). Among them, the high-dose tanshinone extract group inhibited the estradiol level to a level close to that of the positive drug group, showing a clear dose-dependent effect.

[0044] Experiment 4: Effect of tail vein injection of total phenolic acids from tanshinone on serum estradiol levels in ovariectomized mice 1. Test Methods Female BALB / c mice (20±5g) purchased from Spefol (Beijing) Biotechnology Co., Ltd. were randomly divided into four groups: a castration control group (model group), a low-dose total phenolic acid group (TSA-a), a medium-dose total phenolic acid group (TSA-b), and a high-dose total phenolic acid group (TSA-c), with eight mice in each group. The lyophilized powder of total phenolic acid prepared in Example 2 was dissolved in 0.5% CMC-Na solution using vortexing and ultrasonic dissolution methods to prepare solutions with concentrations of 1, 5, and 10 mg / ml. All mice in each group underwent bilateral ovariectomy and were allowed one week of postoperative recovery to eliminate residual estrogen. After the recovery period, mice in each experimental group were administered the following drugs via tail vein injection for 7 consecutive days: the low-dose group received 2 mg / kg / day of total phenolic acid solution, the medium-dose group received 10 mg / kg / day, the high-dose group received 50 mg / kg / day, and the castration control group received an equal volume of 0.9% NaCl solution. The administration frequency was once daily. Blood was collected from the canthus of each mouse before administration and 24 hours after the last administration. The blood samples were centrifuged at 3000 rpm for 15 min, and the supernatant was used to determine the estradiol content in the mice using a mouse estradiol enzyme-linked immunosorbent assay kit (ELISA, purchased from Beijing Mairuida Technology Co., Ltd.).

[0045] 2. Test Results like Figure 5 As shown, compared with the castration control group, all three doses of total phenolic acids injected via the tail vein in the experimental groups reduced estradiol levels in ovariectomized mice. Furthermore, the reduction in serum estradiol levels gradually increased with increasing doses of total phenolic acids, exhibiting a clear dose-dependent effect. These experimental results demonstrate that the total phenolic acids from tanshinone of this invention, when injected via the tail vein in mice, effectively block the conversion of androgens to estrogens and possess significant aromatase inhibitory activity.

[0046] Experimental Example 5: Effect of oral administration of total phenolic acids from Tanshinone on serum estradiol levels in ovariectomized mice 1. Test Methods Female BALB / c mice (20±5g) purchased from Spifort (Beijing) Biotechnology Co., Ltd. were randomly divided into five groups: a castration control group (model group, control), a positive control group (letrozole), a low-dose total phenolic acid group (TSA-a), a medium-dose total phenolic acid group (TSA-b), and a high-dose total phenolic acid group (TSA-c), with eight mice in each group. The lyophilized powder of total phenolic acid prepared in Example 2 was mixed with 0.5% CMC-Na solution as a solvent, and the total phenolic acid was dispersed in the solvent using vortexing and ultrasonic dissolution methods to prepare a total phenolic acid solution. All mice in each experimental group underwent bilateral ovariectomy and recovered for one week post-surgery to eliminate residual estrogen. After the recovery period, mice in each experimental group were administered drugs by gavage for 28 consecutive days as follows: the positive control group received letrozole at a dose of 0.38 mg / kg / day; the low-dose group received total phenolic acid solution of tanshinone at a dose of 10 mg / kg / day; the medium-dose group received total phenolic acid solution of tanshinone at a dose of 50 mg / kg / day; the high-dose group received total phenolic acid solution of tanshinone at a dose of 100 mg / kg / day; and the castration control group received an equal volume of 0.1% CMC-Na solution. The administration frequency was once daily. Blood was collected from the canthus of each mouse before administration and 24 hours after the last administration. The blood samples were centrifuged at 3000 rpm for 15 min, and the supernatant was used to determine the estradiol content in the mice using a mouse estradiol enzyme-linked immunosorbent assay kit (ELISA, purchased from Beijing Mindray Technology Co., Ltd.).

[0047] 2. Test Results As attached Figure 6 As shown, all three groups administered total phenolic acids via gavage reduced estradiol levels in ovariectomized mice to varying degrees, with the TSA-b group (50 mg / kg) showing the greatest decrease. These results indicate that the inhibitory effect of total phenolic acids on estradiol production increased with increasing dosage (from 10 mg / kg / day to 50 mg / kg / day). However, surprisingly, the inhibitory effect decreased with further increases in the concentration of total phenolic acids (from 50 mg / kg / day to 100 mg / kg / day). In conclusion, gavage administration of the total phenolic acids of this invention to mice effectively blocked the conversion of androgens to estrogens and exhibited significant aromatase inhibitory activity.

[0048] In summary, the extract of *Salvia miltiorrhiza* or total phenolic acids of *Salvia miltiorrhiza* in this invention can significantly inhibit aromatase activity and reduce serum estradiol levels in ovariectomized mice. As a key rate-limiting enzyme in the conversion of androgens to estrogens, the inhibition of aromatase activity can directly lead to a reduction in estrogen production in vivo, thereby decreasing the activation of downstream signaling mediated by estrogen receptors.

[0049] Estrogen plays a crucial regulatory role in the proliferation and pathological changes of various tissues. In particular, its abnormally elevated levels are considered a significant driver of disease development and progression in estrogen receptor-positive breast cancer, endometriosis, uterine fibroids, and breast hyperplasia. Therefore, based on the inhibitory effect of tanshinone extract or total phenolic acids on aromatase and the pharmacodynamic evidence of its ability to reduce circulating estradiol levels, it is expected to have potential preventative and therapeutic value for various estrogen-dependent diseases.

[0050] Although specific embodiments of the invention have been described, those skilled in the art will recognize that various changes and modifications can be made to the invention without departing from its scope or spirit. Therefore, the invention is intended to cover all such changes and modifications falling within the scope of the appended claims and their equivalents.

Claims

1. Application of Tanshinone extract in the preparation of aromatase inhibitors.

2. The application according to claim 1, characterized in that, The preparation method of the tanshinone extract is as follows: (1) Weigh out the prescribed amount of Danshen medicinal material, pulverize it and sieve it to obtain medicinal material powder; (2) Add extraction solvent to the medicinal powder obtained in step (1) and extract by reflux to obtain Danshen extract; (3) The extract obtained in step (2) is concentrated and dried to obtain the extract of tanshinone.

3. The application according to claim 1, characterized in that, The tanshinone extract is total phenolic acids from tanshinone.

4. The use of a pharmaceutical composition in the preparation of an aromatase inhibitor, characterized in that, The pharmaceutical composition comprises the tanshinone extract according to any one of claims 1-3.

5. The application according to claim 4, characterized in that, The dosage form of the pharmaceutical composition is selected from oral liquid, tablets, granules, capsules or injections.

6. The application according to claim 3 or 4, characterized in that, The pharmaceutical composition further comprises other aromatase inhibitors.

7. The use of the tanshinone extract according to any one of claims 1-3 or the pharmaceutical composition according to any one of claims 4-6 in the preparation of a medicament for the prevention or treatment of estrogen-dependent diseases.

8. The application according to claim 7, characterized in that, The estrogen-dependent diseases mentioned are selected from breast cancer, breast hyperplasia, endometrial cancer, endometrial hyperplasia, endometriosis, uterine leiomyomas, or gynecomastia.

9. The application according to claim 8, characterized in that, The breast cancer mentioned refers to estrogen receptor-positive breast cancer in postmenopausal women.

10. The application according to any one of claims 7-9, characterized in that, The tanshinone extract is used to treat or prevent estrogen-dependent diseases by inhibiting aromatase activity.

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