CN1730018A - Pharmaceutical composition for treating cardiovascular and cerebrovascular diseases and preparing process and application thereof - Google Patents

Abstract

The invention provides a pharmaceutical composition for treating cardiovascular diseases, which is a medicament prepared by containing total flavonoids of seabuckthorn and seabuckthorn oil as active components. The invention also provides a preparation method of the pharmaceutical composition. The total flavonoids and seabuckthorn oil of the pharmaceutical raw materials of the present invention have played a synergistic effect, strengthened the main effect, overcome side effects, achieved high efficiency, low toxicity, safety, and controllable goals, and are used for treating coronary heart disease, angina pectoris, and hyperlipidemia Etc., the drug is uniformly dispersed, dissolved and released rapidly, and is easy to transport, store, and carry, and is suitable for patients to take, providing a new option for clinical practice.

Description

A pharmaceutical composition for treating cardiovascular diseases, its preparation method and application

technical field

The invention relates to a pharmaceutical composition for treating cardiovascular diseases, specifically, a pharmaceutical composition prepared by combining effective parts extracted from seabuckthorn as raw materials, and belongs to the field of medicine.

Background technique

Cardiovascular disease is a huge burden on global health care and health resources. According to the latest World Health Report released in 2000, 17 million people die from cardiovascular diseases in the world every year, and by 2020, cardiovascular deaths will increase by 50% to as high as 25 million. At present, Western medicine is mainly for symptomatic treatment. Although traditional Chinese medicine has the characteristics of treating both symptoms and root causes, most of them are compound preparations. Due to the constraints of preparation technology and quality standards, it is difficult to be accepted by the international market.

Seabuckthorn is a medicinal material with the characteristics of traditional Chinese medicine. It has a long history of cross-application of traditional Chinese medicine, Tibetan medicine and Mongolian medicine. Since the 8th century AD, seabuckthorn has been used in famous medical classics such as "Yuewang Yaozhen", "Sibu Medical Dictionary" and "Jingzhu Materia Medica". The records of treating diseases are now included in the 2005 edition of "Chinese Pharmacopoeia".

Xindakang Tablets (recorded in the fourteenth volume of standard traditional Chinese medicine preparations issued by the Ministry) are composed of single Chinese medicine extracts of Hippophae rhamnoides L. (seabuckthorn total flavonoids), which have the effect of removing blood stasis and dredging the veins. Clinical studies on 5,000 cases have proved that Xindakang Tablets can improve heart function by 86.9%, improve the total effective rate of ischemic electrocardiogram by 71.8%, treat ischemic heart disease with an effective rate of 81.4%, and simply relieve 15.7%. The effective rate reaches 97.1%, which is superior to western medicine Xiaoxintong, and is considered to be a promising medicine for treating ischemic heart disease by treating both symptoms and root causes. Xindakang Tablets have been used clinically for a long time with positive curative effect. They are mainly used for the treatment of coronary heart disease and angina pectoris. They can improve blood circulation of ischemic myocardium, increase myocardial blood flow, reduce myocardial oxygen consumption, eliminate oxygen free radicals, and improve Oxygen resistance, and can promote and improve the collateral circulation of the myocardium; at the same time, it can reduce blood lipids, prevent blood cell aggregation, reduce the permeability and fragility of capillaries, and is safe and effective for the treatment and prevention of various cardiovascular diseases. Toxic and side effects, worthy of further clinical application. However, the existing Xindakang Tablets are composed of a single active part of total flavonoids of seabuckthorn, and the basic research on its pharmacodynamic substances is relatively lacking. Quantitative analysis only uses spectrophotometry to determine total flavonoids. The research level of quality standards is not high, and the disintegration time is relatively long. The onset is slow, the taking time is long and the number of times is more and other disadvantages.

Seabuckthorn oil is an effective ingredient in seabuckthorn, which contains linoleic acid, carotene, carotenoids, vitamin E, etc. It can reduce blood fat, soften blood vessels, expand blood vessels, and have myocardial and vasodilation effects, ensuring the nutrition and nutrition of the heart muscle itself. The surrounding coronary arteries supply blood and oxygen, reducing the chance of angina pectoris.

At present, there is no relevant report on the combined use of seabuckthorn total flavonoids and seabuckthorn oil in the treatment of cardiovascular diseases.

Contents of the invention

The technical solution of the present invention is to provide a pharmaceutical composition for treating cardiovascular diseases, which is a pharmaceutical composition prepared by combining effective parts extracted from seabuckthorn as raw materials. Another technical solution of the present invention is to provide the drug Methods of preparation and uses of the compositions.

The invention provides a pharmaceutical composition for treating cardiovascular diseases, which is a medicament prepared from raw materials containing the following weight ratios:

Sea buckthorn total flavonoids 1-3 parts, sea buckthorn oil 1-99 parts.

Further, it is a medicament prepared from the following raw materials in weight ratio:

Sea buckthorn total flavonoids 1-3 parts, sea buckthorn oil 1-99 parts.

Furthermore, it is a medicament prepared from the following raw materials in weight ratio:

Sea buckthorn total flavonoids 1-3 parts, sea buckthorn oil 1-9 parts.

Furthermore, it is a medicament prepared from the following raw materials in weight ratio:

Sea buckthorn total flavonoids 3 parts, sea buckthorn oil 7 parts.

Wherein, the total flavonoids contained in the total flavonoids of seabuckthorn are more than 10% by weight calculated as isorhamnetin, and the total fatty acids contained in the seabuckthorn oil are calculated by linoleic acid with a weight percentage of 70% above.

The pharmaceutical composition of the invention is a medicament prepared from total flavonoids of seabuckthorn and seabuckthorn oil as active ingredients, plus pharmaceutically acceptable auxiliary materials or auxiliary ingredients.

Wherein, the medicament is: soft capsule, capsule, tablet, pill, granule, injection.

The present invention also provides a method for preparing the pharmaceutical composition, which comprises the steps of:

a. Take seabuckthorn fruits or seeds or the mixture of fruits and seeds and pulverize them, extract by solvent reflux extraction, supercritical fluid extraction or press centrifugation, and collect seabuckthorn oil;

Seabuckthorn oil is a mixture of seabuckthorn fruit oil or seabuckthorn seed oil or seabuckthorn fruit oil and seabuckthorn seed oil in a certain proportion.

b. Take seabuckthorn fruit and pulverize it, heat and reflux with more than 85% ethanol to extract, concentrate to a thick paste, precipitate with water, filter, degrease the filter cake with petroleum ether, dry the precipitate under reduced pressure, and pulverize to obtain total flavonoids of seabuckthorn; or

Take seabuckthorn medicinal material with n-hexane or solid-phase extraction to degrease, add 80% ethanol to reflux, recover ethanol, add diatomaceous earth to mix well, pump and wash, evaporate to dryness, add ethyl acetate for reflux extraction, recover ethyl acetate, degrease, and obtain Sea buckthorn total flavonoids;

c, get the seabuckthorn total flavonoids and seabuckthorn oil prepared in steps a and b, and prepare pharmaceutically commonly used medicaments according to the following weight ratio:

Sea buckthorn total flavonoids 1-3 parts, sea buckthorn oil 1-99 parts.

The invention also provides the application of the pharmaceutical composition in the preparation of medicine for treating coronary heart disease angina pectoris.

The compound of flavonoids and fat oil, which is prepared according to a certain proportion of medicinal raw materials, has better anti-myocardial ischemia drug effect than "Xindakang Tablet" which uses seabuckthorn total flavonoids as raw materials alone. It also provides the best compatibility ratio of seabuckthorn oil and total flavonoids of seabuckthorn. Seabuckthorn oil and total flavonoids of seabuckthorn have played a synergistic effect. The medicine of the present invention strengthens the main effect, overcomes side effects, integrates and regulates as one, and achieves high efficiency and low cost. Toxicity, safety, and controllable goals, so as to significantly improve the curative effect of Xindakang preparations, and at the same time improve the preparation process and quality control level, improve the pharmacological effects and clinical efficacy of seabuckthorn, and reduce the impurity content at the same time, and can be made into various This preparation is used to treat coronary heart disease, angina pectoris, hyperlipidemia, etc. The drug is uniformly dispersed, dissolved and released quickly, and is easy to transport, store, and carry, and is suitable for patients to take. It provides a new choice for clinical practice.

Apparently, according to the above content of the present invention, according to common technical knowledge and conventional means in this field, without departing from the above basic technical idea of the present invention, other various forms of modification, replacement or change can also be made.

The above-mentioned content of the present invention will be further described in detail below through specific implementation in the form of examples. However, this should not be construed as limiting the scope of the above-mentioned subject matter of the present invention to the following examples. All technologies realized based on the above contents of the present invention belong to the scope of the present invention.

Detailed ways

The extraction of embodiment 1 pharmaceutical raw material seabuckthorn oil of the present invention

Extraction method 1 of seabuckthorn: take seabuckthorn fruit or seeds or a mixture of fruit and seeds, grind them, extract by solvent method under reflux, use n-heptane, heat and reflux extract twice, each time for 4 hours, recover the solvent, and collect seabuckthorn oil;

Seabuckthorn extraction method 2: take seabuckthorn fruit or seeds or a mixture of fruit and seeds, crush them, extract with supercritical CO2 fluid, and collect seabuckthorn oil through a primary separator; wherein, the extraction pressure is 40MPa, the extraction temperature is 65°C, and the extraction The flow rate is 18L/h, the primary separation pressure is 6.5MPa, and the primary separation temperature is 36°C.

Extraction method three of seabuckthorn: squeeze and centrifuge extraction:

Take seabuckthorn fruit or seeds or a mixture of fruits and seeds, squeeze and extract by centrifugation, and mechanically press to obtain seabuckthorn crude oil, separate by centrifugation, and collect seabuckthorn oil;

Embodiment 2 The preparation of total flavonoids of seabuckthorn raw materials of the present invention

Preparation process one:

Take seabuckthorn and grind it into a coarse powder, heat and reflux extraction with more than 85% ethanol three times, each time for 2 hours, filter, combine the filtrate, recover ethanol, concentrate to a thick paste, precipitate with water, filter, and degrease the filter cake with petroleum ether, The precipitate was dried under reduced pressure and crushed. (The total flavonoid content of seabuckthorn is more than 10%)

Preparation process two:

After degreasing the seabuckthorn medicinal material with n-hexane or solid phase extraction (SPE), reflux extraction with 10 times 80% ethanol twice, the recovery rate is ethanol, add 1/4 of the medicinal material with diatomaceous earth, mix well, wash with 20 times the amount of boiling water, and evaporate to dryness , 8 times the amount of ethyl acetate reflux extraction twice, recovery of ethyl acetate, degreasing a small amount of petroleum ether, that is. (The total flavonoid content of seabuckthorn is more than 50%)

The preparation of embodiment 3 drug soft capsules of the present invention

1. Preparation prescription

Raw materials: total flavonoids of seabuckthorn 150g, seabuckthorn oil 350g

Accessories: beeswax 20g

A total of 1000 soft capsules were made from the above (0.52g per capsule)

2. Preparation method

The above two flavors are added with beeswax, ground and mixed in a colloid mill, and compressed into 1000 soft capsules to obtain the final product.

Soft capsules have the following characteristics and are the most suitable dosage form for this product:

1. Neat, beautiful, easy to swallow, light to carry and easy to use.

2. The relative dosage is accurate and the precision is high.

3. Acetaloflavone is insoluble in water, no binder and pressure are needed to prepare soft capsules, basically it is dissolved or fine powder is dispersed in the content, and once disintegrated, it will be released together, and the effect is rapid, which is significantly faster than that of tablets And absorb better.

4. Seabuckthorn oil contains easily oxidized ingredients, which are prepared into soft capsules with sunscreen added to protect the medicine from moisture and the effects of oxygen and light in the air, thereby improving its stability.

5. The prescription of this product contains a high amount of seabuckthorn oil, so it is not easy to be made into a solid drug. It can be made into a soft capsule, which can make up for the shortage of other solid dosage forms.

To sum up, because this product contains water-insoluble solid drug and liquid drug, the dosage of liquid drug is relatively high, and the thick liquid made of two-phase suspension is suitable as soft capsule. This product is made into soft capsules. On the basis of using acetaloflavone, seabuckthorn oil is also used in combination to make up for the shortcomings of hard capsules and tablets in clinical application, such as slow onset and weak effects. The shell can disintegrate quickly, and is convenient for transportation, storage and carrying, and is suitable for patients to take.

The preparation of embodiment 4 pharmaceutical solid preparations of the present invention

①Seabuckthorn oil solidification: Add other preparations to make seabuckthorn oil into solid powder, such as adding starch to absorb and make oil powder

Add gelatin to solidify the package to make oil microcapsules

Add beeswax to solidify, make oily solid, pulverize

Add cyclodextrin for solidification, absorption, crushing, etc.

②Drug mixing: Mix the solid powder of seabuckthorn oil and total flavonoids of seabuckthorn evenly, add appropriate preparation auxiliary materials, and make solid preparations such as tablets, capsules, granules, pills, suspensions, powders, and drop pills

The preparation of embodiment 5 pharmaceutical liquid preparations of the present invention

① To solve the suspension stability of seabuckthorn flavonoids, use the following methods alone or in combination: add oil-soluble surfactants to make microemulsions; add thickening suspensions, ultrafine grinding and wettability, etc.

②Drug mixing: add preparation auxiliary materials to make suspension microemulsion, oil-soluble liquid, etc.

Embodiment 6 Preparation of drug granules of the present invention

Weigh the raw materials prepared in Example 1 and Example 2: 300 g of total flavonoids of seabuckthorn and 700 g of seabuckthorn oil, mix them, add starch, granulate, granulate, and obtain granules.

The preparation of embodiment 7 pharmaceutical tablet of the present invention

Weigh the raw materials prepared in Example 1 and Example 2: 300 g of total flavonoids of seabuckthorn, 700 g of seabuckthorn oil, mix, add starch, granulate, granulate, add magnesium stearate, compress into tablets, to obtain tablets.

The preparation of embodiment 8 medicament oral liquid of the present invention

Weigh the raw materials prepared in Example 1 and Example 2: 300 g of total flavonoids of seabuckthorn and 700 g of seabuckthorn oil, add suspending agent, flavoring agent, and mix with a colloid mill to obtain an oral liquid.

Embodiment 9 Preparation of drug granules of the present invention

Weigh the raw materials prepared in Example 1 and Example 2: 10 g of total flavonoids of seabuckthorn, 990 g of seabuckthorn oil, mix, add starch, granulate, granulate, and obtain granules.

The preparation of embodiment 10 pharmaceutical tablet of the present invention

Take by weighing the raw materials prepared in Example 1 and Example 2: total flavonoids of seabuckthorn 30g, seabuckthorn oil 990g, mix, add starch, granulate, granulate, add magnesium stearate, compress into tablets, to obtain tablets.

Embodiment 11 Preparation of pharmaceutical oral suspension of the present invention

Weigh the raw materials prepared in Example 1 and Example 2: 300 g of total flavonoids of seabuckthorn and 300 g of seabuckthorn oil, add suspending agent, corrective agent, and mix with a colloid mill to obtain an oral suspension.

Embodiment 12 Preparation of pharmaceutical oral suspension of the present invention

Weigh the raw materials prepared in Example 1 and Example 2: 300 g of total flavonoids of seabuckthorn and 900 g of seabuckthorn oil, add suspending agent, corrective agent, and mix in a colloid mill to obtain an oral suspension.

The above-mentioned preparations are all prepared according to the conventional preparation process and selecting conventional auxiliary materials.

By comparison, the stability of solid formulations is better than that of liquid formulations

Embodiment 13 Drug quality control of the present invention

In the present invention, the total flavonoids of seabuckthorn should meet the relevant regulations under the national drug standard-acetaloflavone (WS-10001-(HD-1311)-2003), and the seabuckthorn oil should meet the industry standard-seabuckthorn oil (HB/QS001-94 , HB/QS002-94) and the relevant regulations under the drug standard of seabuckthorn oil raw materials.

Total Fatty Acid Content of Seabuckthorn Oil

Total fatty acid content of seabuckthorn oil (national standard: GB/T17376 preparation of fatty acid methyl esters of animal and vegetable oils, GB/T17377 gas chromatography analysis of fatty acid methyl esters of animal and vegetable oils, gas chromatography; national standard: GB/T15688-1995 in animal and vegetable oils The determination of insoluble impurity content, weight method) is more than 70%, and the results are shown in Table 1.

Relative content of total fatty acids: according to GB/T17376 and GB/T17377, gas chromatography is used for determination, and the relative content is calculated by the peak area normalization method (the content of the peak area normalization method is calculated as 100%).

Relative content of linoleic acid: according to GB/T17376, GB/T17377, gas chromatography determination, the relative content of linoleic acid peak area in the peak area normalization method.

Linoleic acid content: according to GB/T17376, GB/T17377, gas chromatography determination, the weight content of linoleic acid in seabuckthorn oil.

Total flavonoid content of seabuckthorn

The total flavonoid content (National Drug Standard: WS-10001-(HD-1311)-2003, acetaloflavone raw material, spectrophotometric method, calculated as dry product, containing total flavonoid aglycone is calculated as isorhamnetin) is 10% Above, the results are shown in Table 2.

Calculation method of effective part content

Content of effective parts (%) = total flavonoid content of seabuckthorn × its proportion fraction + total fatty acid content of seabuckthorn oil × its proportion fraction × 100

overall scaled score

The average content of the active parts of the medicine of the present invention is calculated to be more than 50%, and the results are shown in Table 3.

In order to verify the effectiveness and safety of the composition of effective parts of seabuckthorn, pharmacodynamic tests and acute toxicity tests were carried out:

Test example 1 Pharmacodynamic screening test of the active part of the drug raw material Hippophae rhamnoides of the present invention:

(1) Effects on electrocardiogram in rats with myocardial ischemia induced by coronary artery ligation

1 Materials and methods

1.1 Animals: Wistar rats, all male, provided by the Experimental Animal Center of Chengdu University of Traditional Chinese Medicine, animal certificate number: Chuan Shi Dong Guan No. 11.

1.2 Drugs and experimental grouping: Seabuckthorn total flavonoids are yellowish-brown dry extract (batch number: 030901, total flavonoid content is 11.5%), provided by Sichuan Meidakang Pharmaceutical Co., Ltd.; seabuckthorn oil is orange-red oily substance (batch number : 031001, with a total fatty acid content of 83.8%), provided by Sichuan Medakang Pharmaceutical Co., Ltd. Rapeseed oil was used to prepare the medicines of each group, and the experimental grouping was as follows, with high and low dose groups respectively. The model control group was given equal volume of rapeseed oil.

Test group S0 S1 S2 S3 S4 S5 S6

Seabuckthorn Oil Ratio 0 50% 60% 70% 80% 90% 100%

Proportion of total flavonoids of seabuckthorn 100% 50% 40% 30% 20% 10% 0

1.3 Reagent: red tetrazolium (TTC), batch number F20030920, produced by China National Pharmaceutical (Group) Shanghai Chemical Reagent Company.

1.4 Instruments: BL-420E four-channel physiological recorder, HX-200 animal ventilator, Chengdu Taimeng Technology Co., Ltd.

2 Experimental methods and results

2.1 Myocardial ischemia modeling: 120 healthy Wistar male rats were randomly divided into groups according to body weight as shown in the table below, administered at different times, administered by gavage for 4 consecutive days, fasting for 12 hours before modeling, and giving After 60 minutes of drug administration, anesthetize with 1.0ml/100g intraperitoneal injection of 10% urethane, fix it on the rat operation board in the supine position, and record its normal electrocardiogram. After the completion, an incision was made on the neck, the tissue was separated, tracheal intubation was performed, and the animal ventilator was connected with a frequency of 50 times/min, a tidal volume of 8-10ml, and an inspiratory-expiratory ratio of 2:1. The thoracotomy was performed at the 4th to 5th intercostal space on the left midclavian line to expose the heart. The pericardium was gently lifted with ophthalmic forceps and torn apart carefully. A needle (3×6, 3/8○) and 000-gauge silk thread were used under the left atrial appendage. The left anterior descending coronary artery was ligated at the coronary sulcus. Then suture the chest cavity, record the electrocardiogram at 5 minutes, 10 minutes, 15 minutes, and 30 minutes after the ligation, compare the changes of the J point (ST segment) before and after the ligation, and use the change value as a statistical index. Animals were sacrificed 4 hours after coronary artery ligation, and the heart was removed by thoracotomy. Rinse it with saline, remove non-cardiac tissue and left and right atrial appendages, absorb water with absorbent paper, weigh it on an electronic balance, which is the weight of the whole ventricle, and cut the heart into 0.1cm thick myocardial slices in parallel from the apex to the bottom of the heart After the myocardial slices were washed with saline, the water was blotted, and the myocardial slices were put into 1% TTC solution. Place in a constant temperature water bath at 37°C and incubate for 5-7 minutes, and shake continuously in the incubating bath to ensure uniform and sufficient staining of the myocardium. Then the myocardium slices were taken out, the excess TTC solution was washed away with normal saline, and after the water was blotted, the stained part of the myocardium was cut off. The unstained part was weighed to obtain the weight of the infarcted myocardium, and the percentage of the infarcted myocardium to the total heart weight was calculated for statistical processing.

The test results are shown in Tables 4 and 5.

The above experimental results show that compared with the model control group, S0, S1, S2, S3, and S4 in the seven ratios can significantly reduce the J-point elevation caused by coronary artery ligation in rats (P<0.05-0.01), suggesting that S0 ~S4 has the effect of protecting myocardial infarction caused by coronary artery ligation, among which S3 (seabuckthorn oil: total flavonoids of seabuckthorn = 70%: 30%) and S4 (seabuckthorn oil: total flavonoids of seabuckthorn = 80%: 20%) have the most obvious effect .

The above results show that among the seven ratios, S0 (seabuckthorn oil: total flavonoids of seabuckthorn=0:100%), S1 (seabuckthorn oil: total flavonoids of seabuckthorn=50%:50%), S2 (seabuckthorn oil: total flavonoids of seabuckthorn=60% %: 40%), S3 (seabuckthorn oil: total flavonoids of seabuckthorn = 70%: 30%), S4 (seabuckthorn oil: total flavonoids of seabuckthorn = 80%: 20%), compared with the model control group, can significantly reduce the size of myocardial infarction (P<0.05-0.01), indicating that S0-S4 can reduce myocardial infarction in rats caused by coronary artery ligation.

(2) Effects on pituitrin-induced myocardial ischemia in rats

1 Materials and methods

1.1 Animals: Wistar rats, half male and half male, provided by the Experimental Animal Center of Chengdu University of Traditional Chinese Medicine, animal certificate number: Chuan Shi Dong Guan No. 11.

1.2 Drugs and grouping: same as above.

1.3 Reagent: Pituitrin, batch number 20030802, produced by Shenyang Jishi Pharmaceutical Co., Ltd., a Sino-US joint venture.

1.4 Instrument: BL-420E four-channel physiological recorder, produced by Chengdu Taimeng Technology Co., Ltd.

2. Experimental method

Pre-screening was carried out before the experiment, and the rats insensitive to vasopressin were eliminated, and 110 qualified ones were randomly divided into groups, administered by intragastric administration for 4 consecutive days, and anesthetized by intraperitoneal injection of 10% urethane half an hour after the last administration. Fixed, subcutaneously inserted ECG electrodes to record lead II ECG. After stable recording for a period of time, pituitary hormone 0.5U/Kg was given by tail vein injection, and the push was completed in 15s. The electrocardiogram was recorded continuously for 20 min. One of the following indications is positive for myocardial ischemia: ①T wave flattening (reducing the height of the original T wave by more than 50%), bidirectional, and inverted; Those without these changes were considered negative. The x2 test was performed with the positive rate of myocardial ischemia in each group.

The test results are shown in Table 6.

The above results show that compared with the model control group, S0, S1, S2, S3, S4, and S5 in the seven ratios can significantly reduce the number of rats with myocardial ischemia caused by intravenous pituitrin (p<0.05) , S0 high (sea buckthorn oil: sea buckthorn total flavonoids=0:100%), S1 high (sea buckthorn oil: sea buckthorn total flavonoids=50%:50%), S3 high (sea buckthorn oil: sea buckthorn total flavonoids=70%:30%) , S4 (seabuckthorn oil: total flavonoids of seabuckthorn = 80%: 20%) has a particularly obvious effect in the high-dose group (p<0.01), indicating that it has a protective effect on myocardial ischemia caused by pituitrin.

(3) Effects on blood lipids in acute hyperlipidemia model mice

1 Experimental materials

1.1 Animals: Kunming mice, half male and half male, provided by the Experimental Animal Center of Chengdu University of Traditional Chinese Medicine, animal certificate number: Chuan Shi Dong Guan No. 11.

1.2 Drugs and grouping: same as above.

1.3 Reagents: Triglyceride (GPO-PAP method detection) kit, batch number 030906, total cholesterol (GOD-CE-PAP method detection) kit, batch number 030910, were purchased from Chengdu Mike Technology Co., Ltd.

2 Experimental methods and results

Modeling method: take 150 mice of 18-22 g, half male and half male, randomly group them into groups, administer medicine continuously for 7 days, and inject 0.5 ml/mouse of 75% egg yolk milk intraperitoneally on the 6th day to form a hyperlipidemia model. After 20 hours, the eyeballs were removed to collect blood, the serum was separated by centrifugation, and the contents of triglyceride (TG) and total cholesterol (TC) in the serum were measured.

The test results are shown in Table 7.

The above test results show that the S2, S3, S4, S5, and S6 groups in the seven kinds of proportions can significantly reduce triglyceride and total cholesterol content in the serum of high-fat rats, wherein S3 (seabuckthorn oil: total flavonoids of seabuckthorn=70% : 30%), S4 (sea buckthorn oil: total flavonoids of sea buckthorn = 80%: 20%), S5 (sea buckthorn oil: total flavonoids of sea buckthorn = 90%: 10%), S6 (sea buckthorn oil: total flavonoids of sea buckthorn = 100%: 0 ) group had the most obvious lipid-lowering effect.

(4) Conclusions of the efficacy screening test of the active part composition

Through the above two anti-myocardial ischemia experiments and blood lipid-lowering experiments, the results showed that the S0, S1, S2, S3, and S4 groups had no effect on rat myocardial ischemia caused by coronary artery ligation and rat myocardial ischemia caused by pituitrin. Ischemia all had obvious protective effects, among which group S3 had the strongest effect; while S2, S3, S4, S5, and S6 had obvious hypolipidemic effects.

The above results show that the total flavonoids of seabuckthorn is the main effective part of anti-myocardial ischemia, and seabuckthorn oil is the main effective part of lowering blood fat. ) has the most prominent comprehensive effect, which not only has obvious anti-myocardial ischemia effect, but also has obvious blood lipid-lowering effect, and fully exerts the effects of the two types of effective parts. The best prescription compatibility ratio of this product is seabuckthorn oil: total flavonoids of seabuckthorn = 7:3.

Pharmacodynamic test of test example 2 pharmaceutical raw materials of the present invention

In order to study the pharmacological effect of the optimal raw material prescription ratio of the effective fraction composition of seabuckthorn in the treatment of ischemic heart disease, the rat model of acute myocardial ischemia was prepared by injecting pituitary hormone and coronary artery ligation. The canine model of myocardial ischemia was prepared by ligation, and the effects of preventive administration on the ST segment of the electrocardiogram, T wave, hemodynamics, myocardial contractility and relaxation performance, and related enzymes in serum (CK, LDH, MDA, SOD, NO) levels of four aspects of the influence, to explore the effective fraction composition of seabuckthorn anti-myocardial ischemia and its mechanism of action; through the observation of hyperlipidemia rats fed with high-fat diet and acute blood stasis model rats , to investigate the blood lipid-regulating effect and the effect on blood rheology of the composition of effective parts of seabuckthorn.

Results: (1) The composition of effective parts of seabuckthorn can significantly improve the ischemic changes of ST segment and T wave of rat electrocardiogram caused by injection of pituitrin and coronary artery ligation (p<0.01)

(2) The composition of effective parts of seabuckthorn can slow down heart rate, lower blood pressure, and improve hemodynamics in rats with acute myocardial ischemia (p<0.01 or p<0.05)

(3) The composition of effective fractions of seabuckthorn has no significant effect on myocardial contractility in rats with acute myocardial ischemia (p>0.05); the composition of effective fractions of seabuckthorn can reduce left ventricular end-diastolic pressure (LVEDP) in rats with acute myocardial ischemia, increasing High left ventricular pressure maximum drop rate (-dp/dtmax), significantly improved myocardial relaxation performance in rats with acute myocardial ischemia (p<0.01 or p<0.05).

(4) The composition of effective parts of seabuckthorn can slow down the heart rate and blood pressure, but has no significant effect on the contractility of the myocardium, indicating that the composition of effective parts of seabuckthorn can reduce the myocardial oxygen consumption in rats with acute myocardial ischemia, and is beneficial to relieve cardiac ischemia. Bloody lesion.

(5) The composition of effective parts of seabuckthorn can significantly reduce the levels of CK, LDH and MDA in the serum of rats with acute myocardial ischemia, increase the activity of SOD and NO, stabilize the structure and function of myocardial cells, and resist ischemic damage (p< 0.01 or p<0.05).

(6) The composition of effective fractions of seabuckthorn can reduce the levels of TC, TG and LDL in the serum of rats with hyperlipidemia, and increase the level of HDL (p<0.01 or p<0.05), indicating that it has a good blood lipid regulating effect.

(7) The composition of effective parts of seabuckthorn can significantly reduce the high, medium and low-shear whole blood viscosity, the high and low-shear whole blood reduced viscosity, and the erythrocyte aggregation index (p<0.01 or p<0.05) of acute blood stasis model rats, showing The composition of effective parts of seabuckthorn can improve blood rheology.

(8) The effect of the composition of effective parts of seabuckthorn on changes of epicardial electrocardiogram ∑-ST and N-ST in dogs with myocardial ischemia caused by coronary artery ligation

Compared with the model control group, the high, medium and low dose groups of the effective part of seabuckthorn composition, the Xindakang tablet control group and the nifedipine group can all significantly resist myocardial ischemia caused by ligation of the coronary arteries. ST and N-ST changes (p<0.05 or p<0.01), while the low-dose group of the effective part composition of seabuckthorn can only resist the ligation of the coronary artery and cause the ligation of the coronary artery to cause myocardial ischemia. Trend of ST change (p>0.05)

(9) The effect of the composition of effective parts of seabuckthorn on the hemodynamic indexes of dogs with myocardial ischemia caused by ligation of coronary arteries

Hemodynamic changes in dogs with acute myocardial ischemia induced by coronary artery ligation: After coronary artery ligation, MAP and HR first slightly decreased, and then returned to the original level or slightly accelerated; LVEDP increased significantly; LVSP, +DP/Dtmax, -DP/ DTmax decreased, and the changes in the above indicators indicated that myocardial ischemia caused impaired cardiac function.

It can be seen from the results that the heart rate HR of each dose group of the effective part composition of seabuckthorn, the Xindakang tablet control group and the nifedipine group decreased to varying degrees at various points after ligation of the coronary artery, and the high, medium and In the low-dose group and the Xindakang tablet control group, the decrease in HR 30 minutes after ligation of the coronary artery was different from that of the model control group (p<0.01); The tablet control group and the nifedipine group had differences (p<0.01 or p<0.05) in MAP at 30 minutes after ligation of the coronary artery with the model control group, while the high, middle and low dose groups of the seabuckthorn effective fraction composition and the heart The Dakang tablet control group has a significant difference with the nifedipine group in the decline rate of MAP (p<0.01); the impact on LVSP: the high, medium and low dose groups of the effective part of seabuckthorn, and the Xindakang tablet control group Compared with the model control group, there were significant differences (p<0.01) between the Nifedipine group and the Nifedipine group at various time points after coronary artery ligation, which was related to their blood pressure lowering effects.

(10) The effect of the composition of effective parts of seabuckthorn on the myocardial contractility and relaxation performance of dogs with myocardial ischemia caused by coronary artery ligation: for +DP/DTmax, each dose group of the composition of effective parts of seabuckthorn, Xindakang tablet control group and Nitrate There was no significant difference between the bendipine group and the model control group after coronary artery ligation (p>0.05). Compared with the model control group, each dose group of the effective part composition of seabuckthorn, Xindakang tablet control group and nifedipine group had no significant effect on Vce40 after ligation (p>0.05); for -DP/Dtmax and LVEDP After ligation, there were significant differences between the high and middle dose groups of the effective parts of seabuckthorn composition, the Xindakang tablet control group and the nifedipine group (p<0.05 or p<0.01) compared with the model control group (p<0.05 or p<0.01), suggesting that the left Ventricular diastolic function, reducing the return of blood, reducing preload.

(11) Effects of seabuckthorn effective fraction composition on myocardial oxygen consumption in dogs with myocardial ischemia caused by coronary artery ligation

The results show that: the composition of effective parts of seabuckthorn, Xindakang tablet control group and nifedipine group can slow down heart rate and blood pressure, but have no effect on myocardial contractility, according to the binomial product of myocardial oxygen consumption "heart rate × blood pressure" It can be inferred that the composition of effective parts of seabuckthorn can reduce myocardial oxygen consumption, improve the energy supply and demand relationship of infarcted myocardium, and relieve ischemic damage.

(12) The effect of the composition of effective parts of seabuckthorn on serum enzymes and myocardial enzyme levels in dogs with myocardial ischemia caused by coronary artery ligation in model group dogs. After coronary artery ligation, serum CK, LDH, and MDA were significantly increased, and NO and SOD were significantly reduced; MDA in myocardial tissue was significantly increased, NO and SOD were significantly decreased (compared with the sham operation group, p<0.01), and the high and middle dose groups of the effective part of seabuckthorn, the Xindakang tablet control group and the nifedipine group were ligated The increase of CK, LDH and MDA in serum and myocardial tissue after coronary artery, the decrease of NO and SOD are significantly different from those in the model group (p<0.05 or p<0.01), and the low dose of the effective part composition of seabuckthorn can resist The trend of this change (p>0.05).

(13) The effect of the composition of effective parts of seabuckthorn on the range of myocardial infarction in dogs with myocardial ischemia caused by coronary artery ligation There is a very significant difference (p<0.01) with the rats in the model group, and there is no significant difference (p>0.05) in the low-dose group of the effective part composition of seabuckthorn.

Conclusion: The above test results show that the composition of effective parts of seabuckthorn has strong myocardial ischemia protection and obvious blood lipid regulation effect, can improve blood rheology, and has a good development and utilization prospect in the treatment of ischemic heart disease.

Test example 3 medicine acute toxicity test of the present invention

Optimum Raw Material Prescription Ratio of Seabuckthorn Effective Parts Composition The maximum concentration and maximum volume of the liquid extract was given to mice once by gavage of 38g/Kg. After 10 minutes of administration, some animals showed restlessness and little movement, and gradually recovered after 30 minutes. normal. There was no abnormality in the urine, stool, color, coat, skin color, nose, eye, and oral secretions of the animals. After the test, the weight of the mice increased, and no animals died. The dosage is 38g of crude drug/Kg (equivalent to 1520 times of the maximum recommended clinical daily dosage for adults).

The best raw material prescription ratio of the effective parts of seabuckthorn. The maximum concentration and maximum volume of the liquid extract were administered to rats once by gavage of 38g/Kg. After 10 minutes of administration, some animals showed static and little movement, and gradually recovered after 30 minutes. normal. There was no abnormality in the urine, stool, color, coat, skin color, nose, eye, and oral secretions of the animals. After the test, the weight of the mice increased, and no animals died. The dosage is 19g of crude drug/Kg (equivalent to 760 times of the maximum recommended clinical daily dosage for adults).

Conclusion: Judging from the dose tested for acute toxicity of this product and animal reactions after administration, this product is a non-obviously toxic drug.

Table 1. Determination results of total fatty acid content in seabuckthorn oil Sample lot number Relative content of linoleic acid (%) Linoleic acid content (mg/g) Total fatty acid content (calculated as linoleic acid, mg/g) 031001021102031101031102040201040202040203041101041102041103 average content 5.33674.20974.07364.97894.52894.56955.07555.73465.75715.22554.9490 45.3633.6734.8234.7236.7235.4236.6944.3142.5342.9538.71 849.96799.82854.77697.34810.79775.14722.88772.68738.74821.93784.41

Table 2. Determination results of total flavonoids in seabuckthorn total flavonoids Sample lot number Total flavonoid content (%) (calculated as isorhamnetin) 020301020302020303030601030602030603040401040402040403040506 11.6111.7514.4912.7214.4912.1615.2312.6312.1612.23

Table 3, composition effective part content measurement result Sea buckthorn oil lot number Seabuckthorn Total Flavonoids Lot Number 1:2 ratio effective part content (%) 1:99 ratio effective part content (%) 021201021202030401030402030701030802031001040201040401040402 020301 60.5357.1960.8550.3657.9255.5552.0655.3853.1258.67 84.2679.3084.7469.1580.3876.8571.6876.6173.2581.49

Table 4. Effects on changes in electrocardiogram J point (mv) of coronary artery ligation model rats (x ± s) group Dose (g/kg) Number of animals (only) normal 5 minutes after piercing 10 minutes after piercing 15 minutes after piercing 30 minutes after piercing Model control group S0 high low S1 high low S2 high low S3 high low S4 high low S5 high low S6 high low Equal volume 0.30.150.30.150.30.150.30.150.30.150.30.150.30.15 888888888888888 0.014±0.020.014±0.020.015±0.020.018±0.020.010±0.020.011±0.020.012±0.030.011±0.020.012±0.030.010±0.020.008±0.020.015±0.020.012 0.020.012±0.020.013±0.02 0.164±0.040.114±0.05*0.116±0.060.114±0.06*0.145±0.050.116±0.040.137±0 050.103±0.02**0.113±0.05*0.115±0.04**0.126±0.060.124±0.080 ±0.100.152±0.090.154±0.10 0.210±0.050.116±0.06*0.118±0.090.120±0.100.132±0.080.116±0.09*0.137±0.100.091±0.06**0.116±0.09*0.121±0.03**0.135±0.100.130.±0.100 154±0.120.155±0.100.157±0.09 0.285±0.080.126±0.08*0.129±0.06*0.125±0.09*0.129±0.160.119±0.11**0.130±0.06**0.111±0.08**0.125±0.09**0.129±0.03**0.135±0.11*0.129 ±0.06**0.165±0.090.162±0.110.169±0.09 0.232±0.080.124±0.09**0.135±0.06*0.116±0.08*0.122±0.09*0.097±0.11**0.125±0.09*0.104±0.08**0.133±0.09**0.146±0.07*0.137±0.09*0.122 0.09*0.146±0.080.160±0.090.165±0.08

Note: Compared with the model control group, *p<0.05, **p<0.01

Table 5. Effect on weight percentage of myocardial infarction area in coronary artery ligation model rats (X±S) group Dose (g/kg) Number of animals (only) Weight percentage of infarction area (%) Model control group S0 high low S1 high low S2 high low S3 high low S4 high low S5 high low S6 high low Equal volume 0.30.150.30.150.30.150.30.150.30.150.30.150.30.15 888888888888888 31.96±5.23 18.37±6.20**20.32±5.86*19.53±3.16**22.00±6.92*18.27±5.30**21.74±7.12*15.64±4.32**20.33±5.46*18.96±4.86**21.01±6.12*28.34 ±4.7824.68±7.4626.52±6.7828.63±5.42

Note: Compared with the model control group, *p<0.05; **p<0.01

Table 6 Effects of pituitrin on acute myocardial ischemia in rats group Dose (g/kg) Number of animals (only) Myocardial ischemia positive number Negative rate of myocardial ischemia Model control group S0 high low S1 high low S2 high low S3 high low S4 high low S5 high low S6 high low Equal volume 0.30.150.30.150.30.150.30.150.30.150.30.150.30.15 101010101010101010101010101010 1045455645346677 0/106/10*5/10**6/10**5/10**5/10**4/10*6/10**5/10**7/10**6/10** 4/10*4/10*3/103/10

Note: Compared with the model control group, *p<0.05; **p<0.01

Table 7 Mouse serum triglyceride content (TG), total cholesterol (TC) content group dose TG content (mmol/L) TC content (mmol/L) Blank control group Model control group S0 high low S1 high low S2 high low S3 high low S4 high low S5 high low S6 high low Equal volume Equal volume 0.30.150.30.150.30.150.30.150.30.150.30.150.30.15 5.64±2.25**16.23±5.9615.74±5.2616.27±5.6012.89±3.3514.72±5.2410.34±4.64*11.46±5.247.26±2.76**8.64±4.59**6.19±2.31**8.02± 3.51**7.94±4.32**8.79±4.61**6.54±3.71**7.26±4.56** 5.69±3.03**13.48±2.6911.64±2.3612.53±2.6210.91±1.7910.83±2.279.17±2.01*9.23±2.35*8.26±2.76**8.88±1.59*7.19±2.31**8.02±1.51 **7.99±2.32**8.79±4.61**7.16±3.25**8.27±2.58**

Note: Compared with the model control group: *p<0.05, *p<0.01

Claims (11)

1. A pharmaceutical composition for treating cardiovascular disease, characterized in that: it is a medicament prepared from raw materials containing the following weight ratios: Sea buckthorn total flavonoids 1-3 parts, sea buckthorn oil 1-99 parts. 2. The pharmaceutical composition for treating cardiovascular diseases according to claim 1, characterized in that it is a medicament prepared from the following raw materials in weight ratio: Sea buckthorn total flavonoids 1-3 parts, sea buckthorn oil 1-99 parts. 3. The pharmaceutical composition for treating cardiovascular diseases according to claim 2, characterized in that it is a medicament prepared from the following raw materials in weight ratio: Sea buckthorn total flavonoids 1-3 parts, sea buckthorn oil 1-9 parts. 4. The pharmaceutical composition for treating cardiovascular diseases according to claim 3, characterized in that it is a medicament prepared from the following raw materials in weight ratio: Sea buckthorn total flavonoids 3 parts, sea buckthorn oil 7 parts. 5. The pharmaceutical composition for treating cardiovascular diseases according to any one of claims 1-4, characterized in that: the total flavonoids contained in the total flavonoids of seabuckthorn are calculated as isorhamnetin, and the weight percentage is 10 % or more, the total fatty acid contained in the seabuckthorn oil is calculated as linoleic acid, and the weight percentage is more than 70%. 6. The pharmaceutical composition for treating cardiovascular diseases according to claim 5, characterized in that it is prepared from total flavonoids of seabuckthorn and seabuckthorn oil as active ingredients, plus pharmaceutically acceptable auxiliary materials or auxiliary ingredients medicine. 7. The pharmaceutical composition for treating cardiovascular diseases according to claim 6, characterized in that: said medicaments are: soft capsules, capsules, tablets, pills, granules, and injections. 8. A method for preparing the pharmaceutical composition for treating cardiovascular diseases as claimed in claim 2, comprising the steps of: a. Take seabuckthorn fruits or seeds or the mixture of fruits and seeds and pulverize them, extract by solvent reflux extraction, supercritical fluid extraction or press centrifugation, and collect seabuckthorn oil; b. Take seabuckthorn fruit and pulverize it, heat and reflux with more than 85% ethanol to extract, concentrate to a thick paste, precipitate with water, filter, degrease the filter cake with petroleum ether, dry the precipitate under reduced pressure, and pulverize to obtain total flavonoids of seabuckthorn; c, get the seabuckthorn total flavonoids and seabuckthorn oil prepared in steps a and b, and prepare pharmaceutically commonly used medicaments according to the following weight ratio: Sea buckthorn total flavonoids 1-3 parts, sea buckthorn oil 1-99 parts. 9. A method for preparing the pharmaceutical composition for treating cardiovascular diseases as claimed in claim 2, comprising the steps of: a. Take seabuckthorn fruits or seeds or the mixture of fruits and seeds and pulverize them, extract by solvent reflux extraction, supercritical fluid extraction or press centrifugation, and collect seabuckthorn oil; b. Take the seabuckthorn fruit and grind it, degrease it with n-hexane or solid phase extraction, add 80% ethanol to reflux, recover ethanol, add diatomaceous earth to mix well, wash and evaporate to dryness, add ethyl acetate to reflux extraction, and recover ethyl acetate , degreasing to obtain total flavonoids of seabuckthorn; c, get the seabuckthorn total flavonoids and seabuckthorn oil prepared in steps a and b, and prepare pharmaceutically commonly used medicaments according to the following weight ratio: Sea buckthorn total flavonoids 1-3 parts, sea buckthorn oil 1-99 parts. 10. The use of the pharmaceutical composition according to any one of claims 1-7 in the preparation of a medicament for treating angina pectoris of coronary heart disease. 11. Use of the pharmaceutical composition according to any one of claims 1-7 in the preparation of a medicament for treating hyperlipidemia.