WO2012094832A1 - Compound chinese medicine extract used for preventing and treating arteriosclerosis and preparation method thereof - Google Patents

Abstract

Disclosed is a compound Chinese medicine extract used for preventing and treating arteriosclerosis, which comprises the following active components: ceryl alcohol, beta-sitosterol, hexacosanic acid, atractylenolide Ⅲ, oleanolic Acid, berberine, jatrorrhizine, coptisine, salvianic acid A, salvianolic acid B, cyclotetracosane, 9,12-linoleic acid, 5,7-dimethoxycoumarin, Specnuezhenide, ginsenoside Rb1 and Rg1, notoginsenoside R1, eucommiol, etc. The said extract is prepared from the raw medicines of danshen root, glossy privet fruit, coptis root, Japanese thistle herb or root, eucommia bark, largehead atractylodes rhizome, panax notoginseng, and bergamot, through mixing effective fractions extracted by organic solvents with different polarity from the total extract of the raw medicine which is extracted by C1-3 alcohol and/or water. Because of elimination of large quantities of noneffective components, the content of the effective components increases largely, and the influence of noneffective components to product processing and formulation quality is decreased. The preparation method is stable and available for mass production with controllable product quality.

Description

 Compound Chinese medicine extract for preventing and treating arteriosclerosis and preparation method thereof

 Technical field

[0001] The present invention relates to the field of traditional Chinese medicine, and in particular to a compound Chinese medicine extract for preventing and treating arteriosclerosis and a preparation method thereof.

 Background technique

[0002] Arteriosclerosis is a common and frequently-occurring disease in the elderly. Atherosclerosis (AS) is the most common and important type of arteriosclerosis, characterized by lipid-like deposition in the intima of the affected arteries, accumulation of complex carbohydrates, followed by fibrous tissue hyperplasia and calcium deposition. There is a lesion in the middle layer of the artery. The disease mainly involves large and medium-sized muscle-elastic arteries. The aorta, coronary arteries and cerebral arteries are common, often leading to serious consequences such as luminal occlusion or wall rupture, which is the main cause of myocardial infarction and cerebral infarction. It affects the large and middle arteries and can cause quite serious consequences. Coronary atherosclerosis, can cause myocardial insufficiency, causing angina and even myocardial infarction, cerebral arteriosclerosis can cause insufficient blood supply to the brain, dizziness, headache. In the later stage of brain atrophy, there may be mental metamorphosis, dementia, etc.; stroke in the cerebral artery or stroke caused by rupture of small aneurysm caused by atherosclerosis (stroke). Can cause loss of consciousness, hemiplegia, aphasia and other serious consequences, renal arteriosclerosis can cause refractory hypertension; intestinal atherosclerosis can cause indigestion, constipation, diarrhea; lower extremity atherosclerosis can cause intermittent claudication, severe extremity Blood necrosis.

[0003] This disease is often accompanied by hypertension, hypercholesterolemia or diabetes. Mental workers are more common, and more men than women. In recent years, the disease has gradually increased in China, which is very harmful to people's health and has become one of the main causes of death in the elderly.

[0004] Due to the economic and social development, the pressure of work and life, the changes in eating habits and lifestyles, and the increase in the incidence of hyperlipidemia and atherosclerosis have become a major threat to human health in contemporary society. It has attracted the attention of the WHO and governments, and is a hot spot in medical research. However, the pathogenesis of atherosclerosis has not yet been fully elucidated, and there is no ideal way to treat atherosclerosis clinically. Although the cause of atherosclerosis is not fully understood, it is known to be closely related to the following factors (susceptibility factors): 1 high blood pressure, 2 hyperlipidemia, 3 smoking, 4 diabetes, 5 obesity. The pathogenesis of atherosclerosis is very much, such as lipid infiltration theory, thrombogenic theory, hemodynamics theory, middle layer smooth muscle cell hyperplasia theory, arterial intimal injury theory, receptor theory, etc., and gradually clear atherosclerosis The pathogenesis of sclerosis is complex and a comprehensive and long process. Recent studies have shown that lipid peroxidation plays an important role in the development of atherosclerosis.

[0005] Although chemical drugs have many drugs for lowering blood lipids or regulating blood rheology, the comprehensive efficacy of preventing and treating atherosclerosis is not satisfactory. Especially because the etiology of atherosclerosis is complicated, and it is accompanied by disorders of lipid metabolism. Hemorheological abnormalities and hyperlipidemia often affect each other. Chemical drugs are usually a special target for lipid metabolism, and pharmacological effects are more specific, improving blood flow. Degeneration and lipid regulation can not be balanced, so a single chemical is difficult to meet the multi-system multi-link pathological complex disease atherosclerosis requires multi-target pharmacological requirements, bringing chemical drugs to treat atherosclerotic disease difficult. In addition, chemical drugs have problems such as adverse reactions and high cost of drugs, which are also the drawbacks of chemicals.

[0006] Clinical practice shows that traditional Chinese medicine in the prevention and treatment of atherosclerosis and cardiovascular and cerebrovascular diseases, because of the overall view of traditional Chinese medicine and dialectical treatment, Chinese medicine has multiple components, can play multi-target, multi-level effect Therefore, the clinical efficacy is exact and has the advantage of being safe and effective.

[0007] Chinese Patent No. 200410051250.4 discloses a drug for treating hyperlipidemia, which is named as 'Fufang Qishu Tiaozhifang' (FTZ), which is made up of Ligustrum lucidum, Atractylodes, Eucommia, Sanqi, Datun, Salvia, and Coptis. The bergamot is composed of Chinese medicine. It has been used for more than 20 years in clinical practice for the treatment of lipid-metabolic diseases such as hyperlipidemia. The experimental study also shows that the extract and the extract also have experimental atherosclerosis in rats. Very good preventive effect.

[0008] Although the compound has a good effect on regulating blood lipids, its effect on the prevention and treatment of atherosclerosis has not been disclosed. Moreover, because it is a compound of traditional Chinese medicine, it has a composition of eight flavors of Chinese medicine, and its chemical composition is complicated, and the active ingredients are not clear. In actual application, the quality of its products is difficult to obtain effective control, which will ultimately affect the stability of the clinical efficacy of the product, and the ineffective components used in the preparation of pharmaceutical preparations are too large, the dosage is large, and the dosage form cannot be diversified, which is not conducive to large-scale production and marketing. Widespread application is limited.

 Summary of the invention

[0009] The object of the present invention is to prevent or treat atherosclerotic compound Chinese medicine, the chemical composition is complex, the active ingredient is unclear, the quality of the product is difficult to obtain effective control, and the number of invalid components is too large. A compound Chinese medicine extract for preventing arteriosclerosis is provided.

[0010] Another object of the present invention to provide a compound Chinese medicine extracts of the above-described production method.

[0011] The object of the present invention is achieved by the following technical solutions:

 A compound Chinese medicine extract for preventing and treating arteriosclerosis, the active ingredients are wax alcohol, β- Sitosterol, n-hexadecanoic acid, atractylenolide III, oleanolic acid, berberine, jatrorrhizine, coptisine, danshensu, salvianolic acid B, cyclotetracosane, 9,12-ten Octadecadienoic acid, 5,7- Dimethoxy coumarin, telomere, ginsenoside Rb1 and Rg1, notoginsenoside R1, eucommia alcohol, and the like.

[0012] The above-mentioned compound Chinese medicine extract is characterized in that the active ingredient laurol, β-sitosterol, n-hexadecanoic acid, atractylenolide III, oleanolic acid, berberine, jatrorrhizine, coptisine, Danshensu, salvianolic acid B, cyclotetracosane, 9,12-octadecadienoic acid, 5,7-dimethoxycoumarin, tetraside, ginsenoside Rb1 and Rg1, Sanqi The weight ratio of saponin R1 and eucommia alcohol is 1 to 5:1 to 6:1 to 4:1 to 4:1 to 8:1 to 8:1 to 5:1 to 5:1 to 6:1 to 5:1 4:1 to 3:1 to 5:1 to 4:1 to 8:1 to 8:1 to 3:1 to 4.

[0013] The preparation method of the above compound Chinese medicine extract comprises the following steps: combining the raw materials Danshen, Ligustrum lucidum, Rhizoma Coptidis, Euphorbia, Eucommia, Atractylodes, Panax notoginseng and bergamot after alcohol extraction and/or water extraction The extract is extracted from the total extract with different polar organic solvents to obtain effective extraction sites of different polarities, and the effective extraction sites are mixed to obtain a compound Chinese herbal extract for preventing and treating arteriosclerosis.

[0014] The specific steps are as follows:

(1) The raw material medicine Panax notoginseng and Ligustrum lucidum were subjected to C _1-3 alcohol extraction to obtain C _1-3 alcohol extract, and the water extracting and concentrating of Datun, Atractylodes Rhizome, Salvia miltiorrhiza, Eucommia, bergamot and Rhizoma Coptidis were carried out to obtain water. Extracting, combining C _1-3 alcohol extract and water extract to obtain total extract;

 (2) extracting the total extract obtained by the step (1) with petroleum ether to obtain an effective extracting site A;

 (3) then extracting the extract remaining after the petroleum ether extraction step (2) with ethyl acetate to obtain an effective extraction site B;

 (4) then extracting the remaining extract after ethyl acetate extraction with n-butanol extraction step (3) to obtain an effective extraction site C Combine the effective parts to obtain a compound Chinese medicine extract for preventing and treating arteriosclerosis.

[0015] The above C _1-3 alcohol is methanol, ethanol or propanol.

[0016] As a preferred embodiment, the C _1-3 alcohol extract is extracted with 30 to 95% by volume of ethanol for 1 to 5 times, preferably 2 to 4 times, and the volume of the C _1-3 alcohol extracted per time is the medicinal material. More than 1.5 times the mass, preferably 6 to 12 times, each extraction time is 5 min to 10 h, preferably 1 to 4 h; the water extraction is 1 to 5 times, preferably 2 to 4 times, each time. The volume of secondary water is 1.5 times or more, preferably 6 to 12 times, and the boiling time is 5 minutes to 10 hours, preferably 1 to 4 hours. The volume of the concentrate after concentration is 0.2 to the mass of the medicine. 5 times, preferably 1 to 2 times.

[0017] In the above preparation method, the method for preparing the effective extraction site A is as follows: adding 0.5 to 10 times, preferably 3 to 5 times the volume of petroleum ether of the extract, extracting 1 to 5 times, preferably 2 to 4 times, combining Extracting, recovering petroleum ether, and drying at low temperature to obtain part A; the effective extraction site B is prepared as follows: adding 0.5 to 15 times, preferably 5 to 12 times the volume of the extracted extract of ethyl acetate, and extracting 1 to 5 times, Preferably, the mixture is extracted 2 to 4 times, and the ethyl acetate is recovered and dried at a low temperature to obtain a portion B. The effective extraction site C is prepared by adding 0.5 to 15 times, preferably 5 to 12 times the volume of the extract. Alcohol, extraction 1 to 5 times, preferably 2 to 4 times, combining the extracts, recovering n-butanol, and drying at a low temperature to obtain a portion C.

[0018] In the compound Chinese medicine prepared by the present invention, the weight ratio of the effective parts A, B and C is from 0 to 1:0 to 2:0 to 3.

[0019] In the present invention, each effective site or a combination thereof can be processed into a tablet, a capsule, a granule, an oral liquid preparation, a pill, and a parenteral in accordance with a pharmaceutically acceptable carrier and a general preparation method. In the form of powder injection, suspension, injection, and the like.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

By the preparation method of the invention, a large number of ineffective chemical components in the traditional Chinese medicine can be removed, and different effective extraction sites can be obtained, which not only retains the characteristics of the traditional Chinese medicine, but also greatly increases the content of the active ingredients in the traditional Chinese medicine, and reduces the ineffective ingredients to the product processing and preparation. The influence of quality makes the effective ingredients of the compound Chinese medicine clear, the preparation process is stable, the product quality is controllable, and it is beneficial to mass production.

 DRAWINGS

[0021] FIG. 1 : Example 2 Fingerprint of compound Chinese herbal extract (FTZCE) analyzed by UHPLC-MS.

 detailed description

[0022] The following examples further explained in conjunction with the present invention, but the embodiment of the present invention do not form any limitation.

 [0023] Example 1 Preparation of Total Extract of Traditional Chinese Medicine Compound

 Will be female, Atractylodes, Eucommia, Panax notoginseng, Datun, Salvia, Coptis, bergamot (weight ratio 1:1:1:2:1:2:1:1) In the case of the Sanqi and Ligustrum lucidum, the two medicines were pulverized into coarse powder, and the extracts were extracted three times with the volume of 12 times, 10 times and 8 times of 60 times of ethanol. The combined extracts are recovered, and the ethanol is recovered and the concentration of the alcohol extract is 1 ml, which is equivalent to 1 g of crude drug; the other herbs in the square are decomposed and extracted three times with water of 12 times, 10 times and 8 times, respectively. h, three times of aqueous extracts were combined, concentrated under vacuum to 1 g of medicinal material / ml extract, and the water extracting part and the previous alcohol extracting part were combined to obtain the total extract of traditional Chinese medicine compound (FTZ).

 Example 2 Preparation of Effective Extraction Site of Compound Total Extract

 Extract once with 5 times volume of petroleum ether from the total extract of the compound, separate the petroleum ether layer; extract it separately in 3 times and 2 times volume. The petroleum ether layer was separated and the petroleum ether extract was combined three times to recover petroleum ether, and then dried under vacuum at 0.08 MPa and 55 ° C to obtain an effective extraction site A;

 The remaining extract of the total extract of petroleum ether is added 12, 8, and 5 respectively. A large amount of ethyl acetate was extracted, and the obtained ethyl acetate extraction site was recovered, and the solvent was recovered, and dried under vacuum at 0.07 MPa and 60 ° C to obtain a compound effective extraction site B;

 The remainder after ethyl acetate extraction is again 10, 8, and 6 A plurality of n-butanol were separately extracted, and the extracts were combined to recover n-butanol, and the extract was vacuum dried at 0.07 MPa and 60 ° C to obtain a compound effective extraction site C.

[0025] According to the total amount of the traditional Chinese medicine compound and the amount of each effective extraction site, the yield of each extract is calculated.

[0026] The yield of the effective extraction site A is between 0.10% and 0.30%.

 The effective extraction site B yield is 0.40% to 1.00%

 The yield of effective extraction site C is between 2.00% and 4.00%.

 According to the effective extraction site A, B and C, the weight ratio of 1: 2:5 is combined with the compound Chinese herbal extract for preventing arteriosclerosis ( FTZCE), the fingerprint obtained by UHPLC-MS analysis is shown in Figure 1, which is characterized by the inclusion of lauryl alcohol, β- Sitosterol, n-hexadecanoic acid, atractylenolide III, oleanolic acid, berberine, jatrorrhizine, coptisine, danshensu, salvianolic acid B, cyclotetracosane, 9,12-ten Octadecadienoic acid, 5,7- Active ingredients such as dimethoxycoumarin, tetraside, ginsenoside Rb1 and Rg1, notoginsenoside R1, and eucommia alcohol.

 [0027] Example 3 Preparation method of compound Chinese medicine extract capsule

 The extract is combined in the following parts by weight:

 Effective extraction site A 1 part

 Effective extraction site B 2 parts

 Effective extraction site C 5 parts

 The extracts in the above ratio, the effective extraction site A and the same amount of starch are uniformly mixed, and then the effective extraction site B Mix and finally mix with the effective extraction site C. The three are mixed uniformly and then dry granulated. After drying under reduced pressure, the whole granules are filled with No. 2 capsules, polished and examined.

[0028] The obtained capsules were determined by HPLC method, and the results of the contents of the respective active ingredients are shown in Table 1:

 Table 1 FTZCE capsule HPLC content determination results

Test items	Content (mg/granule)	Test items	Content (mg/granule)
Oleanolic acid	2.02	Alcohol	1.06
Female glucoside	0.67	Berberine	2.57
Salvian acid	1.58	Coptidis	1.01
N-hexadecanoic acid	0.55	Atractylenolide III	0.91
Ginsenoside Rb1	2.41	Ginsenoside Rg1	2.50
5,7-dimethoxycoumarin	0.65	Eucommia alcohol	0.85

 Example 4 Effect of Chinese herbal compound extract on experimental arteriosclerosis in rats

 Experimental animals: SD rats, SPF grade, male, weighing 200 g ± 20 kg.

[0029] Method: Rats were adaptively reared for one week and randomly divided into normal group, model group, FTZCE (prepared by method 2) low dose group, FTZCE high dose group, extraction sites A, B, C and FTZ ( Example 1 Method Preparation) Group. Among them, 10 normal control groups were fed with basal feed and 80 models were made. The model group and each drug-administered group received a single intraperitoneal injection of vitamin D3 600,000 units/kg, while feeding high-fat feed and 10 mL/kg per day. A rat model of arteriosclerosis was prepared by fat milk (fat milk self-matching: 25 g of cholesterol per 500 mL of fat milk, 5 g of sodium cholate, 2.5 g of propylthiouracil, and 75 g of lard). The low-dose and high-dose groups of the composition were administered with FTZCE 30 mg/(kg·d) and 60 mg/(kg·d) per day, respectively. The extract site A 10 mg/(kg · d) and the extract site B 40 mg/ (kg · d), extraction site C 120mg / (kg · d) and FTZ 500 mg / (kg · d) suspension, the normal group and the model group were given the same amount of distilled water per day. Once a day for 8 weeks, blood lipids, TNFα, IL-1, IL-6, IL-10 and NF-kB were measured by venous venous plexus 60 minutes after the last administration, and the abdominal aorta was measured. Determination of blood rheology indicators. The rats were sacrificed and the aorta, heart and liver were immersed in 10% formalin. The arterial, cardiac and hepatic lesions were observed by paraffin embedding, sectioning and HE staining. The thickness of the media and the radius of the vascular lumen were calculated. Ratio, immunohistochemical analysis using SABC method to observe the positive expression of NF-κB in the arterial membrane, and to observe the lesions of the arteries. The experimental results are shown in Table 2 and Table 3.

[0030] Table 2 Effect of FTZCE on blood lipids in experimental rats (n=10 mean±standard deviation, unit: mmol/L)

Group	Dosage mg/kg	TC	TG	LDL-C
Normal control group	/	1.26±0.21**	0.98±0.16**	0.67±0.25**
Model group	/	2.86±0.27	1.84±0.28	1.26±0.38
FTZ	500	1.64±0.25*	1.23±0.16*	0.71±0.08*
Extraction site A	10	1.58±0.22**	1.29±0.21*	0.70±0.13*
Extraction site B	40	1.62±0.31*	1.30±0.19*	0.65±0.28**
Extraction site C	120	1.55±0.26**	1.48±0.33	0.81±0.24*
FTZCE low dose group	30	1.52±0.20**	1.28±0.22*	0.66±0.23**
FTZCE high dose group	60	1.50±0.34*	1.23±0.25*	0.68±0.21**

 Note: Compared with the high fat model group, * P < 0.05, ** P < 0.01.

[0031] Table 2 shows that the compound extract and extract composition of the present invention is prepared by high-fat feeding plus a single intraperitoneal injection of vitamin D ₃ 600,000 units/kg, while perfusing 10 mL/kg of fat milk per day. The rat arteriosclerosis model has a significant regulatory effect on abnormal lipid metabolism in rats, which can significantly reduce the levels of cholesterol and low-density lipoprotein in the model rats, increase the ratio of high-density lipoprotein to low-density lipoprotein, and inhibit the model. Thickening of the membrane in the arterial wall of rats.

[0032] The concentration of TNF α, IL-1, IL-6 and NF-kB in the model group was significantly increased, and the concentration of IL-10 was significantly decreased; the composition of the compound extract, A, B, C and extract was high or low. The dose can significantly reduce the concentration of inflammatory factors TNFα, IL-1, IL-6, IL-10 and NF-kB, increase the concentration of IL-10; inhibit the overexpression of arterial NF-KB, see Table 3.

[0033] Table 3 Effect of FTZCE on blood TNFα, IL-1, IL-6, IL-10 and NF-kB in experimental rats (n=10 mean±standard deviation, unit: р mol/L)

Group	Dosage mg/kg	TNFa	IL1	IL6	NF-KB	IL10
Normal control group	/	1.06±0.11**	1.36±0.21**	1.26±0.25**	1.18±0.16**	1.67±0.25**
Model group	/	2.36±0.27	2.28±0.28	2.76±0.28	2.51±0.38	1.01±0.38
FTZ	500	1.84±0.25*	1.84±0.25*	1.96±0.35*	1.23±0.16*	1.51±0.08*
Extraction site A	10	1.78±0.22**	1.88±0.24**	1.92±0.28**	1.79±0.23*	1.35±0.13*
Extraction site B	40	1.92±0.31*	1.82±0.33*	1.86±0.35*	1.90±0.39*	1.36±0.25**
Extraction site C	120	1.85±0.26*	1.95±0.21**	1.95±0.29**	1.78±0.33	1.28±0.23*
FTZCE	30	1.82±0.20**	1.52±0.20**	1.72±0.32**	1.58±0.29*	1.45±0.24**
FTZCE	60	1.60±0.34**	1.60±0.34*	1.60±0.38*	1.41±0.23*	1.61±0.23**

 Note: Compared with the model group, * P < 0.05, ** P < 0.01.

[0034] Effect on rat blood rheology indicators: Compound extract and extract compositions can reduce whole blood and plasma viscosity and inhibit platelet aggregation, see Table 4.

[0035] Table 4. Effect of compound extracts and their compositions on hemorheology in experimental hyperlipidemic rats (n = 10)

Drug (mg/kg)	FTZCC 30	FTZCC 60	FTZ 500	FTZ -A 10	FTZ-B 40	FTZ -C 120	Control group
Whole blood viscosity, high cut (200/s)	5.85±0.75 ★★	5.25±0.3 ★★	5.6±0.42 ★★	5.416±0.34 ★★	5.48±0.42 ★★	5.76±0.41 ★★	7.01±0.45
Whole blood viscosity, low cut (3/s)	11.6±2.62 ★★	11.12±2.03 ★★	12.34±2.56 ★★	11.34±2.16 ★★	12.04±2.51 ★★	12.73±2.16 ★★	15.18±2.64
Plasma viscosity (mpa/s)	1.62±0.35 ★★	1.56±0.47 ★★	1.60±0.42 ★★	1.64±0.42 ★	1.70±0.45 ★★	1.69±0.38 ★★	2.23±0.39
Fibrinogen ( g/L )	3.86±0.33 ★★	3.27±0.38 ★★	4.19±0.90 ★★	4.11±0.90 ★★	3.99±0.90 ★★	4.16±0.88 ★★	4.47±0.36
Normal platelet aggregation %	31.8±7.5 ★★	25.25±4.3 ★★	25.6±4.2 ★★	25.19±4.1 ★★	27.6±4.1 ★★	28.6±4.6 ★★	37.0±4.5
ADP induces platelet aggregation %	61.6±6.6 ★★	53.12±6.3 ★★★	62.4±5.6 ★★	60.4±5.2 ★★	68.6±5.3 ★★	67.4±5.8 ★★	100.±6.6 ★★

Note: Compared with the control group, ^★ P <0.01, ^▲ P <0.05.

[0036] Table 5 Effect of compound extract and its composition on the ratio of medial thickness and lumen radius of abdominal aorta in rats with arteriosclerosis (n=6)

Group	Dosage mg/kg	Medium film thickness / lumen radius ( μ m)
Normal control group	/	0. 17 ± 0. 01
Model group	/	0. 35 ± 0. 04
FTZ	500	0. 19 ± 0. 03 *
Extraction site A	10	0. 25 ± 0. 03 *
Extraction site B	40	0. 26 ± 0. 02 **
Extraction site C	120	0. 27 ± 0. 03 *
FTZCE	30	0. 21 ± 0. 03 **
FTZCE	60	0. 18 ± 0. 02 **

 Note: Compared with the high fat model group, * P < 0.05, ** P < 0.01.

[0037] The results of pathological observation showed that the vascular structure of the normal group was clear, the inner elastic membrane was intact, and the smooth muscle cells of the middle membrane were arranged regularly, parallel to the inner elastic membrane. In the model group, atherosclerotic plaque appeared in the aorta of the rats. The intravascular elastic membrane damage was broken, the middle membrane was obviously irregularly thickened, the ratio of the thickness of the medial membrane to the radius of the vascular lumen was significantly increased, and the smooth muscle cells were disorderly arranged. Intimal migration tendency, smooth muscle fibers were significantly disordered; myocardial tissue lipid deposition was severe, and a large number of red stained granules appeared in the cytoplasm of myocardial cells in the lesion; liver oil staining was strongly positive. The intravascular elastic membrane rupture of the rats in each treatment group was reduced compared with the model group, and some of the tensile and flattening were flattened, and the smooth muscle fibers of the middle membrane were slightly disordered. The low and high doses of the compound extract and its composition reduced the thickness of the medium film to varying degrees. / The effect of intravascular lumen radius ratio was significantly different between the low dose group and the model group (P < 0.01). The results are shown in Table 5. Lipid deposition in myocardial tissue and liver tissue of rats in each treatment group was also significantly alleviated.

 Example 5 Effect of Oxidized Low Density Lipoprotein on Vascular Smooth Muscle Cell Proliferation

Methods: Refer to Wang Daosheng's patch method (Wang Daosheng, arterial smooth muscle cell culture method, see: Xu Shuyun, editor of pharmacological experimental methodology, Beijing: People's Medical Publishing House, 2001: 573), 6-week-old male SD rats were killed by decapitation The thoracic aorta media was taken and cultured in DMEM medium containing 10% fetal bovine serum in a patch culture method, and cultured in a 37 ° C, 5% CO ₂ incubator for primary culture of rat vascular smooth muscle cells. . The third to fifth generation passages of vascular smooth muscle cells were selected for oxidized low density lipoprotein (Ox-LDL)-induced vascular smooth muscle cell proliferation experiments, and different concentrations of Chinese herbal compound extracts and their compositions were observed for oxidized low density lipoprotein (Ox- LDL) induces the effects of vascular smooth muscle cell proliferation.

[0039] For cell proliferation evaluation, refer to the method established by Selzman CH et al. (Selzman CH, et a. Class II cytokine receptor ligands inhibit human vascular smooth muscle proliferation. Surgery, 1998, 124: 318), SD rat vascular smooth muscle cells with 0.25 % trypsinized and plated at a density of 5000 cells/well in 96-well plates. After 8 hours, the cells were replaced with serum-free, glutamic acid-containing DMEM medium, and serum-free conditions were maintained for 48 h to obtain synchronized cell growth stop, and then replaced with DMEM medium containing 5% FBS and the corresponding experimental agent, Ox-LDL. The final concentration was 100 μg/ml, and incubated with vascular smooth muscle cells for 24 h. Each group was repeated 6 times, each group was 6 wells. The control group was DMEM medium containing 5% FBS, and the cells were treated for 24 h. The proliferation rate was determined by applying the CellTiter 96 Assay MTS/PES kit.

[0040] The specific method is MTS / PES 20 μ l added to the corresponding wells, 96-well plates were incubated in a 5% CO ₂ , 37 ° C incubator for 90 min, and then in the Mithras LB 940 multi-function microplate reader at 490 nm The absorbance was measured and the weight per well was 3 times, and the proliferation rate of the cells was expressed by absorbance. The results are shown in Table 6.

[0041] Table 6 Effect of extract composition on proliferation of rat vascular smooth muscle cells stimulated by Ox-LDL (n=6)

Group	Drug concentration (μg/ml)	VSMCs value-added rate	VSMCs value-added rate
		No Ox-LDL cell OD value	Ox-LDL cell OD value
normal	0	0.839±0.137	1.482±0.213
FTZ	10	0.832±0.128	1.091±0.132**
FTZ	50	0.823±0.136	0.913±0.140**
FTZCE	2	0.829±0.129	1.011±0.135**
FTZCE	5	0.831±0.131	0.993±0.141**
FTZCE	10	0.810±0.157	0.839±0.137**
FTZCE	50	0.811±0.137	0.819±0.162**

 Note: Compared with the Ox-LDL group alone, **P<0.05.

[0042] The results showed that 100 μg/ml of Ox-LDL significantly stimulated the proliferation of rat vascular smooth muscle cells (P<0.05, see Table 6). The FTZ total extract and extract composition (FTZCE) alone had no effect on the cell proliferation rate of vascular smooth muscle cells. Compared with the control group, the FTZ total extract and extract composition (FTZCE) induced Ox-LDL in rats. Proliferation of vascular smooth muscle cells is significantly inhibited.

[0043]

Experimental studies have shown that the pharmaceutical composition obtained by the method of the invention can also effectively lower serum total cholesterol, low density lipoprotein cholesterol content, lower the content of cholesterol in tissues (such as liver and heart), and lower serum triglyceride levels, Platelet aggregation is also inhibited. In addition, the product of the invention can also inhibit the proliferation of smooth muscle, prevent thickening of the inner membrane and damage of endothelial inflammation, prevent thrombosis, inhibit and reverse the damage of arteriosclerosis, and thereby improve the overall health of the human body.

Claims (10)

1. A compound Chinese medicine extract for preventing and treating arteriosclerosis, the active ingredients are wax alcohol, β-sitosterol, n-hexadecanoic acid, atractylenolide III, oleanolic acid, berberine, jatrorrhizine, coptisine, Danshensu, salvianolic acid B, cyclotetracosane, 9,12-octadecadienoic acid, 5,7-dimethoxycoumarin, tetraside, ginsenoside Rb1 and Rg1, Sanqi Saponin R1, eucommia alcohol.
2. The compound Chinese herbal medicine extract according to claim 1, characterized in that the active ingredient laurol, β-sitosterol, n-hexadecanoic acid, atractylenolide III, oleanolic acid, berberine, jatrorrhizine, berberine Alkali, Danshensu, salvianolic acid B, cyclotetracosane, 9,12-octadecadienoic acid, 5,7-dimethoxycoumarin, telomere, ginsenoside Rb1 and Rg1, The weight ratio of notoginsenoside R1 and eucommia alcohol is 1 to 5:1 to 6:1 to 4:1 to 4:1 to 8:1 to 8:1 to 5:1 to 5:1 to 6:1 to 5: 1 to 4:1 to 3:1 to 5:1 to 4:1 to 8:1 to 8:1 to 3:1 to 4.
3. The method for preparing a compound Chinese medicine extract according to claim 1, comprising the steps of:

   (1) extracting the raw materials Danshen, Ligustrum lucidum, Rhizoma Coptidis, Euphorbia, Eucommia, Atractylodes, Sanqi and Bergamot with water and/or C _1-3 alcohol to obtain total extract;

   (2) extracting the total extract obtained in the step (1) with different polar organic solvents to obtain effective extraction sites of different polarities, and mixing the effective extraction sites to obtain a compound Chinese herbal extract for preventing and treating arteriosclerosis.
4. The preparation method according to claim 3, comprising the steps of:

   (1) The raw material medicine Panax notoginseng and Ligustrum lucidum were subjected to C _1-3 alcohol extraction to obtain C _1-3 alcohol extract, and the water extracting and concentrating of Datun, Atractylodes Rhizome, Salvia miltiorrhiza, Eucommia, bergamot and Rhizoma Coptidis were carried out to obtain water. Extracting, combining C _1-3 alcohol extract and water extract to obtain total extract;

   (2) extracting the total extract obtained by the step (1) with petroleum ether to obtain an effective extracting site A;

   (3) then extracting the extract remaining after the petroleum ether extraction step (2) with ethyl acetate to obtain an effective extraction site B;

   (4) Then extracting the remaining extract after ethyl acetate extraction in the n-butanol extraction step (3) to obtain an effective extraction site C, and combining the effective extraction sites to obtain a compound Chinese herbal extract for preventing and treating arteriosclerosis.
5. The preparation method according to claim 4, wherein the C _1-3 alcohol extraction means extracting 1 to 5 times with 30 to 95% by volume of C _1-3 alcohol, and extracting C _1-3 alcohol each time. The volume is more than 1.5 times the mass of the medicinal material, and the extraction time is 5 min to 5 h.
6. The preparation method according to claim 4, characterized in that the water extraction is decocted 1 to 5 times with water, and the volume of each water is 1 to 15 times the mass of the medicinal material, and the boiling time is 5 minutes to 10 hours each time. The concentrated concentrate has a volume of 0.2 to 5 times the mass of the medicinal material.
7. The preparation method according to claim 4, wherein the effective extraction site A is prepared by adding 0.5 to 15 times the volume of the extracted petroleum ether, extracting 1 to 5 times, combining the extracts, and recovering petroleum ether. The low-temperature drying results in an effective extraction site A.
8. The preparation method according to claim 4, wherein the effective extraction site B is prepared by adding 0.5 to 15 times the volume of the extracted extract of ethyl acetate, extracting 1 to 5 times, combining the extracts, and recovering the ethyl acetate. The low-temperature drying results in an effective extraction site B.
9.  The preparation method according to claim 4, wherein the effective extraction site C is prepared by adding 0.5 to 15 times the volume of the extract to n-butanol, extracting 1 to 5 times, combining the extracts, and recovering n-butanol. The low-temperature drying results in an effective extraction site C.
10. The preparation method according to claim 4, wherein the weight ratio of the effective extraction sites A, B and C is 0 to 1:0 to 2:0 to 3.

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