阅读说明：本技术 一种治疗功能失调性子宫出血的头顶一颗珠有效部位及制备方法 (Effective part of berberis capitata for treating dysfunctional uterine bleeding and preparation method ) 是由 杨竹雅 丁雄 刘录 马晓霞 王晓珍 王钺涵 谭文红 周志宏 于 2020-12-31 设计创作，主要内容包括：本发明属于医药技术领域,涉及一种中药提取物,具体涉及一种治疗功能失调性子宫出血的头顶一颗珠有效部位及制备方法。本方案的药物是由头顶一颗珠药材经提取加工制成的甾体皂苷有效部位,甾体皂苷有效部位包括三种以一定比例组成的甾体皂苷。本方案的药物可通过口服发挥其止血、缩宫作用,达到治疗功能失调性子宫出血目的,毒副作用少,具有良好的开发应用价值。(The invention belongs to the technical field of medicines, relates to a traditional Chinese medicine extract, and particularly relates to an effective part of Aconitum capitatum for treating dysfunctional uterine bleeding and a preparation method thereof. The medicine of the scheme is a steroid saponin effective part prepared by extracting and processing a medicinal material of berberis kunmingensis, and the steroid saponin effective part comprises three steroid saponins formed according to a certain proportion. The medicine of the scheme can play the roles of stopping bleeding and contracting uterus through oral administration, achieves the aim of treating dysfunctional uterine bleeding, has little toxic and side effects and has good development and application values.)

1. An effective part of barberry lily bulb for treating dysfunctional uterine bleeding is characterized by comprising a steroid saponin effective part extracted from barberry lily bulb medicinal materials; the effective parts of the steroid saponin comprise pennogenin-3-alpha-L-rhamnopyranosyl (1 → 4) -beta-D-glucoside, pennogenin-3-O-alpha-L-rhamnopyranosyl (1 → 4) - [ alpha-L-rhamnopyranosyl (1 → 2) ] -beta-D-glucoside and pennogenin-3-alpha-L-rhamnopyranosyl- (1 → 4) -O-alpha-L-rhamnopyranosyl- [ O-alpha-L-rhamnopyranosyl- (1 → 2) ] -O-beta-D-glucoside.

2. The effective fraction of Aconitum capitatum for treating dysfunctional uterine bleeding according to claim 1, the method is characterized in that the mass ratio of the pennogenin-3-alpha-L-rhamnopyranosyl (1 → 4) -beta-D-glucoside to the pennogenin-3-O-alpha-L-rhamnopyranosyl (1 → 4) - [ alpha-L-rhamnopyranosyl (1 → 2) ] -beta-D-glucoside to the pennogenin-3-alpha-L-rhamnopyranosyl- (1 → 4) -O-alpha-L-rhamnopyranosyl- [ O-alpha-L-rhamnopyranosyl- (1 → 2) ] -O-beta-D-glucoside is 70-80; 10-20; 5 to 10.

3. The effective fraction of barberry lily what is used for treating dysfunctional uterine bleeding according to claim 2, wherein the dosage of the effective fraction of steroid saponin is 30mg three times a day, and the dosage form of the effective fraction of steroid saponin is any pharmaceutically acceptable dosage form.

4. A preparation method of an effective fraction of Tibet barberry root for treating dysfunctional uterine bleeding is characterized in that the effective fraction of steroid saponin is obtained by sequentially carrying out alcohol extraction, petroleum ether extraction, macroporous resin purification and alcohol precipitation on the medicinal material of Tibet barberry root.

5. The method for preparing the effective fraction of the first barberry lily bulb for the treatment of dysfunctional uterine bleeding according to claim 4, wherein in the alcohol extraction step, the first barberry lily bulb medicinal material is pulverized to obtain coarse powder, and the coarse powder is subjected to cold-leaching extraction by using ethanol to obtain an extraction solution; and concentrating the extracting solution to obtain an extract I.

6. The method for preparing an effective fraction of Plectranthus Amboinicus for treating dysfunctional uterine bleeding according to claim 5, wherein said coarse powder is extracted by cold extraction with ethanol three times in said alcohol extraction step.

7. The method for preparing the effective fraction of Achillea alpina for treating dysfunctional uterine bleeding according to claim 6, wherein the extract I is dispersed in water to obtain an extract dispersion liquid in the petroleum ether extraction step; extracting the extract dispersion with petroleum ether and collecting the water layer.

8. The method for preparing the effective fraction of Achillea alpina for treating dysfunctional uterine bleeding according to claim 7, wherein the macroporous resin purification step comprises adsorbing the water layer by using macroporous resin, performing gradient elution by using ethanol and water, collecting the eluate at a volume ratio of ethanol to water of 70:30, and concentrating the eluate to obtain extract II.

9. The method for preparing an effective fraction of Achillea alpina for treating dysfunctional uterine bleeding according to claim 8, wherein the volume percentage of ethanol is increased from 0 to 90% by gradient elution with ethanol and water in the macroporous resin purification step.

10. The method for preparing the effective fraction of the first barberry lily bulb for the treatment of dysfunctional uterine bleeding according to claim 7, wherein in the alcohol precipitation step, the extract II is dissolved in an ethanol solution under a heating condition, and after cooling, the precipitate is filtered and dried to obtain the effective fraction of the steroid saponin.

Technical Field

The invention belongs to the technical field of medicines, relates to a traditional Chinese medicine extract, and particularly relates to an effective part of Aconitum capitatum for treating dysfunctional uterine bleeding and a preparation method thereof.

Background

Dysfunctional uterine bleeding, abbreviated as dysfunctional uterine bleeding, is caused by neuroendocrine dyscrasia, but not by systemic or female genital tract organic diseases such as pregnancy, endometrial tumor, infection or hematopathy. It usually occurs in adolescence or perimenopause, and is mostly anovulatory dysfunctional uterine bleeding, which is a common gynecological disease. Dysfunctional uterine bleeding can have various clinical manifestations, mainly manifested as abnormal menstrual cycle frequency, regularity, menstrual period length, menstrual bleeding volume, often complicated anemia, infection, etc. At present, the hemostasis treatment is mainly carried out clinically by taking hormone medicines such as compound gestodene and the like, and the surgery treatment such as uterine curettage, hysterectomy and the like is carried out on serious patients. The ideal medicine for treating dysfunctional uterine bleeding can achieve the treatment purpose by controlling and treating clinical symptoms, regulating the uterine function and taking hormone medicines for no long time, and the medicine with the functions still is an important medicine research and development direction in the field. In view of the harm of dysfunctional uterine bleeding and the demand for the development of related drugs, the development of a specific drug for treating dysfunctional uterine bleeding with high efficiency and little side effect is urgently needed.

Disclosure of Invention

The invention aims to provide an effective part of Aconitum capitatum for treating dysfunctional uterine bleeding, which is used for solving the technical problem that a specific medicine for treating the dysfunctional uterine bleeding with high efficiency and small side effect is lacked in the prior art. According to the scheme, the medicinal material of the first berberis from Yunnan province is used as a raw material, the effective part of the steroid saponin for treating functional uterine bleeding is prepared, and the treatment effect of the medicinal material of the first berberis on the functional uterine bleeding is further improved. The effective part of the steroid saponin can achieve the effects of stopping bleeding, contracting uterus and resisting inflammation through oral administration or injection, and has good treatment effect on functional uterine bleeding.

The technical scheme of the invention is as follows:

an effective fraction of radix Berberidis kunmingensis for treating dysfunctional uterine bleeding comprises steroid saponin effective fraction extracted from radix Berberidis kunmingensis; the effective parts of the steroid saponin comprise pennogenin-3-alpha-L-rhamnopyranosyl (1 → 4) -beta-D-glucoside, pennogenin-3-O-alpha-L-rhamnopyranosyl (1 → 4) - [ alpha-L-rhamnopyranosyl (1 → 2) ] -beta-D-glucoside and pennogenin-3-alpha-L-rhamnopyranosyl- (1 → 4) -O-alpha-L-rhamnopyranosyl- [ O-alpha-L-rhamnopyranosyl- (1 → 2) ] -O-beta-D-glucoside.

The principle of the technical scheme is as follows: radix seu caulis Parthenocissi Tricuspidatae, also called rhizoma Trillii Tschonossii, rhizoma lion Hemsleyae Macrospermae, and radix seu caulis Parthenocissi Tricuspidatae, are dried rhizome of Trillium tschonoskii Maxim, distributed in Tibet, Yunnan, Guizhou, Sichuan, Hunan, Hubei, etc., and have effects of tranquilizing, promoting blood circulation, stopping bleeding, removing toxic substance, etc., and can be used for treating hypertension, neurasthenia, giddiness, headache, traumatic injury, traumatic hemorrhage, etc. The main functional component of the parietal barberry is steroid saponin, wherein the steroid saponin comprises pennogenin and diosgenin which are connected with different glycosyl chains to form different components, and the varieties are various. At present, it is not clear which of the steroid saponins are main functional components, which limits the further development of the root of common cephalanoplos medicinal material.

The first barberry medicine is generally used as a hemostatic and blood coagulation medicine, and the medicine is not applied to the treatment of dysfunctional uterine bleeding. Treatment of dysfunctional uterine bleeding requires not only that the drug have a hemostatic function to relieve clinical symptoms, but also that it be capable of regulating uterine function (e.g., uterine contractility) as well as human hormone levels. The inventor applies the medicinal material of berberis capitata to the treatment of dysfunctional uterine bleeding for the first time, and the medicinal material shows good effects of stopping bleeding and promoting uterine contraction.

How to obtain a finely processed product with better treatment and prevention effects on functional uterine bleeding by using a medicinal material of berberis poiretii is still a problem to be solved in the field. In order to improve the treatment effect of the medicinal material of the common cephalanoplos herb, the inventor conducts a great deal of research on the functional components of the common cephalanoplos herb, and finds that the extract (steroid saponin) containing three steroid saponins extracted from the common cephalanoplos herb has good effect of treating dysfunctional uterine bleeding.

The beneficial effect of this scheme lies in:

(1) the effective part of the steroid saponin is extracted from the medicinal material of the Aconitum kusnezoffii, and the pennogenin which is naturally sourced has good hemostatic and anti-inflammatory effects and small toxic and side effects.

(2) The effective part of the steroid saponin has the effects of stopping bleeding, contracting uterus and resisting inflammation, and has good treatment effect on dysfunctional uterine bleeding.

(3) The effective part of the steroid saponin has low toxicity and high safety when being orally administrated. When the daily dosage of the mouse is 1mg/kg, no obvious toxic reaction of the animal is seen. The mice are subjected to continuous 14-day administration experiments, and toxicity changes related to administration, such as general conditions, liver and kidney functions and main organ coefficients, are not found.

In conclusion, the scheme provides the effective part of the Aconitum capitatum for treating dysfunctional uterine bleeding, which has the advantages of simple and convenient preparation process, small medicament dosage, less toxic and side effects, low price and economy and good development and application values.

Further, the mass ratio of the pennogenin-3-alpha-L-rhamnopyranosyl (1 → 4) -beta-D-glucoside, the pennogenin-3-O-alpha-L-rhamnopyranosyl (1 → 4) - [ alpha-L-rhamnopyranosyl (1 → 2) ] -beta-D-glucoside to the pennogenin-3-alpha-L-rhamnopyranosyl- (1 → 4) -O-alpha-L-rhamnopyranosyl- [ O-alpha-L-rhamnopyranosyl- (1 → 2) ] -O-beta-D-glucoside is 70-80; 10-20; 5 to 10.

By adopting the technical scheme, through a large amount of researches on the medicinal material of the root of Chinese barberry produced in different Yunnan provinces, the inventor finds that the ratio of three steroid saponins in the effective part of the steroid saponin obtained by adopting the extraction method is 70-80; 10-20; 5 to 10. In addition, the inventor verifies through experiments that the mixture formed by the three steroid saponins in the proportion has good hemostatic and blood coagulation effects and good treatment effects on dysfunctional uterine bleeding.

Furthermore, the dosage of the effective part of the steroid saponin is 30mg three times a day, and the dosage form of the effective part of the steroid saponin is any pharmaceutically acceptable dosage form.

By adopting the technical scheme, the effective part of the steroid saponin can achieve the effects of stopping bleeding, contracting uterus and resisting inflammation through various modes such as oral administration or injection, and the like, and has good treatment effect on functional uterine bleeding.

Further, a preparation method of the effective component of the Achillea alpina for treating dysfunctional uterine bleeding comprises the steps of sequentially carrying out alcohol extraction, petroleum ether extraction, macroporous resin purification and alcohol precipitation on the Achillea alpina medicinal material to obtain the steroid saponin effective component.

By adopting the technical scheme, firstly, alcohol is used as a solvent to soak and extract the medicinal material of the berberis capitata, so that a large amount of steroid saponins in the medicinal material are leached from the medicinal material; then removing part of non-polar impurities by using a petroleum ether extraction method; further separating and enriching the target components by macroporous resin adsorption and gradient elution; finally, further removing impurities by alcohol dissolving and precipitating, and purifying the effective part of the steroid saponin. The effective part of the steroid saponin extracted by the scheme contains three functional components, namely pennogenin-3-alpha-L-rhamnopyranosyl (1 → 4) -beta-D-glucoside, pennogenin-3-O-alpha-L-rhamnopyranosyl (1 → 4) - [ alpha-L-rhamnopyranosyl (1 → 2) ] -beta-D-glucoside and pennogenin-3-alpha-L-rhamnopyranosyl- (1 → 4) -O-alpha-L-rhamnopyranosyl- [ O-alpha-L-rhamnopyranosyl- (1 → 2) ] -O-beta-D-glucoside. The three functional components have combined action, and have remarkable curative effects on blood coagulation, uterine contraction promotion and dysfunctional uterine bleeding treatment.

Further, in the alcohol extraction step, crushing the medicinal material of the common cephalanoplos herb to obtain coarse powder, and performing cold-leaching extraction on the coarse powder by using ethanol to obtain an extracting solution; and concentrating the extracting solution to obtain an extract I.

By adopting the technical scheme, the steroid saponin in the first-root-of-Chinese-barberry medicinal material can be fully extracted by cold soaking and extraction of ethanol, and a prerequisite condition is created for later separation, enrichment and purification.

Further, in the alcohol extraction step, the coarse powder is subjected to cold extraction three times using ethanol.

By adopting the technical scheme, the effective components in the medicinal materials are fully extracted by multiple times of extraction.

Further, in the petroleum ether extraction step, dispersing the extract I in water to obtain an extract dispersion liquid; extracting the extract dispersion with petroleum ether and collecting the water layer.

By adopting the technical scheme, petroleum ether is used for extraction, and nonpolar ingredients in the extract dispersion liquid are removed, so that the target functional ingredients in the water layer can be enriched.

Further, in the step of purifying by macroporous resin, adsorbing the water layer by using macroporous adsorption resin, performing gradient elution by using ethanol and water, collecting eluent when the volume ratio of the ethanol to the water is 70:30, and concentrating the eluent to obtain an extract II.

By adopting the technical scheme, the macroporous resin adsorption and gradient elution modes are adopted, and the eluent containing the target component can be obtained when the ethanol aqueous solution with the volume fraction of 70% is used for elution. According to subsequent efficacy experimental studies, the eluent of the part has very remarkable effect of promoting blood coagulation, and the blood coagulation promoting effect of the eluent obtained by using the eluent with other proportion is poor. The extraction and purification method of the scheme effectively obtains the effective part of the steroid saponin with stronger drug effect, and plays a role in promoting the curative effect of the medicinal material of the berberis pubescens on the functional uterine bleeding.

Further, in the macroporous resin purification step, when gradient elution is performed using ethanol and water, the volume percentage of ethanol increases from 0 to 90%.

By adopting the technical scheme, impurities are gradually separated through gradient elution, and target functional components are enriched and purified. The effective part of the steroid saponin containing three types of pennogenin glucoside can be obtained by the macroporous resin adsorption and gradient elution method, and the effective part of the steroid saponin has excellent effects of stopping bleeding, contracting uterus and resisting inflammation and has good treatment effect on functional uterine bleeding.

Further, in the alcohol precipitation step, dissolving the extract II in an ethanol solution under the heating condition, cooling, filtering to obtain a precipitate, and drying the precipitate to obtain the steroid saponin effective part.

By adopting the technical scheme, through the processes of heating dissolution and cooling precipitation of the ethanol, impurities which can be dissolved in the ethanol are removed, and the effective part of the steroid saponin is further purified.

Detailed Description

Example 1

Taking 50kg of the medicinal material of the common cephalanoplos herb, crushing the medicinal material of the common cephalanoplos herb into coarse powder, respectively carrying out cold-leaching extraction for three times (48h, 24h and 24h) by using 10 times of ethanol (the coarse powder is ethanol-1 kg and is 10L), combining extracting solutions, and carrying out reduced pressure concentration to obtain an extract I (8.55kg) (the relative density is controlled to be 1.25-1.30, and the relative density is specifically 1.26 in the embodiment); dissolving extract I with 5 times of water (1 kg: 5L, 43L) under stirring to obtain extract dispersion. Extracting with petroleum ether of the same volume as the extract dispersion for 3 times, adsorbing a water layer with macroporous adsorption resin (D101), performing gradient elution with ethanol and water (0: 100-70: 30), eluting with water for 130L, eluting with ethanol (10%, 30%, 50%, 70%, 90%) of different concentrations respectively, eluting with 130L of each concentration, collecting 70% ethanol elution part to obtain an eluate containing a target component, and concentrating the eluate under reduced pressure to obtain an extract II (4.8kg) (the relative density is controlled to be 1.25-1.30, and the relative density is specifically 1.28 in the embodiment). Adding 5 times of 30% ethanol (1 kg: 5L) into extract II, heating to dissolve, standing for 24 hr, filtering, and drying the precipitate at 70 deg.C to obtain white powder (2.6kg) to obtain effective components of steroid saponin.

Through Nuclear Magnetic Resonance (NMR) and mass spectrum detection (MS), the chemical components of the effective part of the steroid saponin are identified as follows: pennogenin-3-alpha-L-rhamnopyranosyl (1 → 4) -beta-D-glucoside (compound A), pennogenin-3-O-alpha-L-rhamnopyranosyl (1 → 4) - [ alpha-L-rhamnopyranosyl (1 → 2) ] -beta-D-glucoside (compound B) and pennogenin-3-alpha-L-rhamnopyranosyl- (1 → 4) -O-alpha-L-rhamnopyranosyl- [ O-alpha-L-rhamnopyranosyl- (1 → 2) ] -O-beta-D-glucoside (compound C). In addition, the mass ratio of the compound A, the compound B and the compound C is 76:16:8 after the effective part of the steroid saponin is analyzed by HPLC.

The spectral data are specifically as follows:

a compound A: white needle crystal. ESI-MS m/z: 737[ M-H]The indicated relative molecular weight is 738. Determining the molecular formula as C by combining carbon spectrum₃₉H₆₂O₁₃。1H-NMR(C5D5N，500MHz)δ：0.67(3H，d，J＝5.8Hz，H-27)，0.93(3H，s，H-18)，1.08(3H，s，H-19)，1.22(3H，d，J＝7.2Hz，H-21)，5.26(1H，brd，J＝5.0Hz，H-6)，1.76(3H，d，J＝6.2Hz，2'-O-Rha-H-6)，4.98(1H，brd，Glc-H-1)，6.37(1H，brd，2'-O-Rha-H-1)。

Compound B: white needle crystal. ESI-MS m/z: 883[ M-H ]]The relative molecular weight is 884. Determining the molecular formula as C by combining carbon spectrum₄₅H₇₂O₁₇。1H-NMR(C5D5N，500MHz)δ：0.67(3H，d，J＝5.3Hz，H-27)，0.95(3H，s，H-18)，1.07(3H，s，H-19)，1.22(3H，d，J＝7.2Hz，H-21)，5.28(1H，brd，J＝4.2Hz，H-6)，1.75(3H，d，J＝6.1Hz，2'-O-Rha-H-6)，1.62(3H，d，J＝6.1Hz，4'-O-Rha-H-6)，4.82(1H，brd，Glc-H-1)，5.85(1H，brd，2'-O-Rha-H-1)，6.39(1H，brd，4'-O-Rha-H-1)。

Compound C: white needle crystal. ESI-MS m/z: 1030 binding of carbon profiling moleculesFormula is C₅₁H₈₂O₂₁。1H-NMR(C5D5N，500MHz)δ：0.68(3H，d，J＝5.4Hz，H-27)，0.95(3H，s，H-18)，1.07(3H，s，H-19)，1.22(3H，d，J＝7.2Hz，H-21)，5.29(1H，brd，J＝3.8Hz，H-6)，1.58(3H，d，J＝5.6Hz，4”-O-Rha-H-6)1.75(3H，d，J＝6.2Hz，2'-O-Rha-H-6)，1.61(3H，d，J＝6.5Hz，4'-O-Rha-H-6)，4.84(1H，brd，Glc-H-1)，5.82(1H，brd，2'-O-Rha-H-1)，6.38(1H，brd，4'-O-Rha-H-1)，7.05(1H，brd，4”-O-Rha-H-1)。

Example 2

By adopting the method of example 1, the effective parts of the steroid saponins are extracted from 18 batches of the medicinal material of the berberis pubescens, and the chemical component composition (mass) proportion of the effective parts of the steroid saponins in different batches is shown in table 1.

Table 1: example 2 preparation of effective part of steroid saponin and test results

Serial number	Origin of medicinal materials	Batch number	Batch (kg)	The component composition ratio (A: B: C)
					1	Yunnan Huaping	Y1701	20	76:16:8
2	Yunnan Huaping	Y1701	20	74:17:9
					3	Yunnan Huaping	Y1801	20	78:15:7
4	Yunnan Huaping	Y1801	20	76:16:8
					5	Yunnan Huaping	Y1802	15	72:19:9
6	Yunnan Huaping	Y1802	15	73:17:10
					7	Guizhou Fenggang	G1703	25	79:16:5
8	Guizhou Fenggang	G1703	25	78:16:6
					9	Guizhou Fenggang	G1704	20	75:15:10
10	Guizhou Fenggang	G1704	20	75:16:9
					11	Guizhou Fenggang	G1803	20	74:18:8
12	Guizhou Fenggang	G1803	20	75:16:9
					13	Enshi in Hunan province	H1705	30	70:20:10
14	Enshi in Hunan province	H1705	30	71:19:10
					15	Enshi in Hunan province	H1804	20	75:17:8
16	Enshi in Hunan province	H1804	20	76:17:7
					17	Enshi in Hunan province	H1805	20	80:10:10
18	Enshi in Hunan province	H1805	20	79:11:10

From the results in the above table, it can be seen that the compound ratio ranges are compound a: 70% -80%; compound B: 10% -20%; compound C: 5 to 10 percent.

Example 3

Pulverizing the effective part of steroid saponin obtained in example 1 into fine powder, weighing 300g, adding 700g starch, mixing, and encapsulating 10000 capsules, each containing 100mg effective part of steroid saponin. The medicine is orally taken by patients with functional uterine bleeding three times a day, 1 granule each time.

Experimental example 1: hemostasis effect of effective part of steroid saponin

1. Sample (I)

Positive control group: GONGXUENING Capsule (Yunnan white drug powder group GmbH, specification 0.13 g/capsule, product batch number ZMA1402) is prepared into 3.38mg/ml with distilled water, and daily dosage is 0.3 ml. A suspension of concentration;

sample high dose group: the sample prepared in the example 1 is added with water to prepare 0.78mg/ml suspension, and the daily dose is 0.3 ml;

sample low dose group: the sample prepared in the example 1 is added with water to prepare 0.39mg/ml suspension, and the daily dose is 0.3 ml;

normal group: physiological saline

Control group 1: taking the water eluent concentrate (prepared into extract according to the method of the example 1) in the example 1, adding water to prepare 0.78mg/ml suspension, and the daily dose is 0.3 ml;

control group 2: taking the 10% ethanol eluate concentrate of example 1 (prepared into extract according to the method of example 1), adding water to obtain 0.78mg/ml suspension, with daily dose of 0.3 ml;

control group 3: collecting 30% ethanol eluate concentrate of example 1 (prepared into extract by the method of example 1), adding water to obtain 0.78mg/ml suspension, with daily dose of 0.3 ml;

control group 4: taking the 50% ethanol eluate concentrate of example 1 (prepared into extract according to the method of example 1), adding water to make into 0.78mg/ml suspension, with daily dose of 0.3 ml;

control group 5: the 90% ethanol eluate concentrate of example 1 (prepared into extract by the method of example 1) was taken, and water was added to make 0.78mg/ml suspension, 0.3ml daily dose.

2. Laboratory animal

Healthy Kunming mouse, body weight 19-23 g. Purchased to the laboratory animal center of Kunming medical university.

3. Test methods and procedures

The test was carried out according to the method and operation of the reference "the experimental study of the hemostatic effect of gynecological hemostatic mixture on functional uterine bleeding" (Shanghai: the study of the university of TCM college in Hubei, see pages 9-10 of the study, Exit and blood coagulation time experiments). The effect of the drug on promoting blood coagulation was tested by this experiment.

4. Test results

The experimental results are shown in table 2, and the results show that the bleeding and blood coagulation time is obviously shortened when the positive control group, the sample low-dose group and the sample high-dose group are compared with the normal control group; compared with the positive control group, the bleeding and blood coagulation time of the sample high-dose group and the sample low-dose group is obviously shortened, wherein the high-dose group has the best effect; the other elution stages had essentially no effect. It is demonstrated that the fraction of the parietal barberry with the best biological efficacy (steroid saponin efficacy fraction) can be obtained by the method of the present act.

Table 2: influence of effective part of steroid saponin on bleeding and blood coagulation time of mouse

Group of	Number of samples (N)	Bleeding time (min)	Blood coagulation time (min)
				Normal group	10	20.37±0.39	4.87±0.65
Positive control group	10	16.76±0.52Δ	3.55±0.34Δ
				Sample Low dose group	10	14.37±0.74Δ*	3.07±0.45Δ*
Sample high dose group	10	12.15±0.68Δ*	2.46±0.28Δ*
				Control group 1	10	19.88±0.34	4.62±0.69
Control group 2	10	20.71±0.36	4.91±0.52
				Control group 3	10	20.03±0.45	4.78±0.48
Control group 4	10	20.61±0.42	4.82±0.58
				Control group 5	10	19.92±0.48	4.93±0.80

Δ P <0.05 compared to normal group; comparison of P <0.05 with control group

Experimental example 2: the effective part of steroid saponin has uterine contraction effect

1. Sample (I)

Positive control group: gongxuening capsule (Yunnan Baiyao group GmbH, 0.13 g/capsule, product batch ZMA1402) is prepared into suspension with concentration of 0.273mg/ml with distilled water. The daily gavage of the rabbits was recorded as 100 ml.

Sample high dose group: adding water into the sample prepared in the example 1 to prepare 0.021mg/ml, wherein the daily gavage dose is 100 ml;

sample low dose group: adding water into the sample prepared in the example 1 to prepare 0.0105mg/ml, wherein the daily gavage dose is 100 ml;

normal group: physiological saline;

control group 1: taking the water eluent concentrate (prepared into extract according to the method of the example 1) in the example 1, adding water to prepare 0.021mg/ml suspension, and taking 100ml of daily dose;

control group 2: taking 10% ethanol eluate concentrate of example 1 (prepared into extract by the method of example 1), adding water to make into 0.021mg/ml suspension, and making into 100ml daily dose;

control group 3: taking the 30% ethanol eluate concentrate of example 1 (prepared into extract by the method of example 1), adding water to make into 0.021mg/ml suspension, and making into 100ml daily dose;

control group 4: taking the 50% ethanol eluate concentrate of example 1 (prepared into extract by the method of example 1), adding water to make into 0.021mg/ml suspension, and making into 100ml daily dose;

control group 5: the concentrate of 90% ethanol eluate of example 1 (prepared into extract by the method of example 1) was added with water to make 0.021mg/ml suspension, and the daily dose was 100 ml.

2. Laboratory animal

Healthy and non-pregnant rabbits with the weight of about 1400-1600 g. Purchased to the laboratory animal center of Kunming medical university.

3. Test methods and procedures

The test was carried out according to the method and operation of the reference "the experimental study of the hemostatic effect of gynecological hemostatic mixture on functional uterine bleeding" (Shanghai: the study of the university of Hubei college of traditional Chinese medicine, pp 13-14, the experiment of uterine suspensory). The effect of the drug on promoting uterine contraction was verified by this experiment.

4. Test results

The experimental results are shown in table 3, and the results show that the positive control group and the sample group have significant contraction effect on the uterus compared with the normal control group; compared with a positive control group, the sample group has more remarkable uterine contraction effect; the other elution stages had essentially no effect.

Table 3: effect of effective part of steroid saponin on rabbit uterine contraction

Group of	Number of samples (N)	Frequency (times/min)	Amplitude (mm)	Physical activity
					Normal group	10	6.00±1.10	2.84±1.14	12.42±7.62
Positive control group	10	7.33±1.75*	3.92±0.66	28.96±9.10**
					Sample Low dose group	10	9.67±2.25**	3.93±0.61	33.25±9.80**
Sample high dose group	10	10.50±2.07**	5.46±0.62**	46.55±9.99**
					Control group 1	10	5.88±1.25	2.97±1.05	11.68±5.87
Control group 2	10	6.40±1.30	2.80±1.08	12.29±6.36
					Control group 3	10	5.93±1.22	2.75±1.26	12.06±7.45
Control group 4	10	6.12±1.01	2.71±1.20	12.39±8.04
					Control group 5	10	6.39±1.04	2.95±1.28	12.90±7.78

P <0.05 compared to normal group; comparison of P <0.01 with normal group

Experimental example 3: safety test of effective part of steroid saponin

The effective part of the steroid saponin in the embodiment 1 is taken and suspended by water, and the ICR mice are gavaged for 14 days continuously with 1mg/kg/day dosage 1 time a day to give samples, without the toxicity change related to the administration of general conditions, liver and kidney functions and main organ coefficients.

The foregoing is merely an example of the present invention and common general knowledge in the art of designing and/or characterizing particular aspects and/or features is not described in any greater detail herein. It should be noted that, for those skilled in the art, without departing from the technical solution of the present invention, several variations and modifications can be made, which should also be regarded as the protection scope of the present invention, and these will not affect the effect of the implementation of the present invention and the practicability of the patent. The scope of the claims of the present application shall be determined by the contents of the claims, and the description of the embodiments and the like in the specification shall be used to explain the contents of the claims.