Composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction, as well as preparation method and application thereof

Abstract

The present invention relates to a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction, as well as its preparation method and application, and belongs to the field of traditional Chinese medicine. The preparation method of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction is as follows: crushing epimedium and then extracting it with ethanol; crushing Corydalis yanhusuo, leucopsis pilosula, ginkgo biloba and salvia miltiorrhiza and then mixing them and extracting them with ethanol; crushing hawthorn and then hydrolyzing them with hydrogen peroxide ultrasonically; adding the alcohol-extracted epimedium residues and the mixed residues to the hawthorn hydrolyzate, adding a composite enzyme for hydrolysis to obtain an enzymatic solution; subjecting the enzymatic solution to microwave-assisted reflux extraction and concentration to obtain a water-extracted clear paste; mixing the epimedium alcohol-extracted clear paste, the mixed alcohol-extracted clear paste and the water-extracted clear paste and filtering them, adding gypenosides to the filtrate to obtain a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction. The composition obtained by the preparation method of the present invention has increased oligosaccharide and polysaccharide contents, improved stability of glycoside components, and improved bioavailability, and has the effects of promoting blood circulation, reducing turbidity, reducing atherosclerotic plaques, preventing the formation of atherosclerotic plaques, and preventing myocardial infarction.

Description

Technical Field

[0001] The present invention relates to the technical field of traditional Chinese medicine preparation, in particular to a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction, and a preparation method and application thereof. Background Art

[0002] Coronary atherosclerotic heart disease (CHD) is a condition characterized by abnormal lipid metabolism, which causes lipids in the blood to deposit on the previously smooth lining of the arteries. These atherosclerotic lipids accumulate in the arterial lining, forming white plaques known as atherosclerotic lesions. The gradual accumulation of these plaques narrows the arterial lumen, obstructing blood flow and leading to ischemia and hypoxia in the heart, resulting in symptoms such as angina, chest pain, tightness, and shortness of breath. Further episodes of angina may lead to myocardial infarction (MI). MI is a severe form of CHD, characterized by acute occlusion of the coronary arteries on top of stenosis, completely cutting off distal blood flow and causing severe ischemia, hypoxia, and even necrosis of parts of the myocardium.

[0003] Coronary heart disease (CHD) has long been a major threat to Chinese patients. In recent years, with changes in lifestyles and improvements in living standards, the incidence of CHD has gradually increased, with a trend of younger patients, seriously impacting people's health. Qi deficiency and blood stasis are common pathogenesis of CHD, and blood stasis is closely related to the formation of coronary atherosclerotic plaques. Activating blood circulation and reducing turbidity are important approaches to prevent and treat myocardial infarction (MI) caused by CHD. Traditional Chinese Medicine emphasizes promoting blood circulation and reducing turbidity to improve blood quality, reduce blood viscosity, and help maintain vascular patency, thereby preventing MI.

[0004] The applicant's prior invention patent CN113730467A discloses a pharmaceutical composition for treating coronary heart disease and its preparation method. The raw materials for the preparation are gypenosides, ginkgo leaves, salvia miltiorrhiza, Corydalis yanhusuo, hawthorn, leucanthemum officinale, and epimedium. The composition obtained by this preparation method contains glycosides, terpenoid lactones, and flavonols as the main active ingredients. The main glycoside components are icariin, 2"-O-galloyl-codonoside, and phloroglucin. However, actual studies have found that icariin has poor water solubility and easily precipitates during storage. Furthermore, icariin is not easily absorbed, resulting in low bioavailability. Furthermore, 2"-O-galloyl-codonoside and phloroglucin have poor stability in aqueous solutions and degrade during storage, resulting in reduced content in oral solutions formed from the composition. Therefore, further research and development of this composition is needed to overcome the problems existing in existing compositions. , in order to further enhance its blood circulation and turbidity reduction effects, reduce the formation of atherosclerotic plaques, and achieve a more effective effect in preventing myocardial infarction. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to provide a method for preparing a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction, so that the composition can overcome the problems of poor stability and low bioavailability of the glycoside active ingredients by increasing the content of oligosaccharides and polysaccharides, and can further reduce the level of plasma low-density lipoprotein cholesterol and remove toxic heavy metal ions in the blood, thereby more effectively achieving the effect of preventing myocardial infarction.

[0006] To solve the above technical problems, the present invention provides a method for preparing a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction. The raw materials of the composition include ginkgo biloba, salvia miltiorrhiza, gypenosides, Corydalis yanhusuo, hawthorn, herba leucanthemums and epimedium. The preparation method of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction comprises the following steps:

[0007] 1) crushing raw material Epimedium, soaking it in an ethanol solution with a concentration greater than 80%, heating and refluxing for extraction, filtering to obtain Epimedium alcohol extract and Epimedium residue; recovering ethanol from the Epimedium alcohol extract to obtain Epimedium clear paste;

[0008] 2) crushing the raw materials Corydalis yanhusuo, gypsophila gracile, ginkgo biloba, and salvia miltiorrhiza, mixing them uniformly, soaking them in a 60-70% ethanol solution, heating and refluxing them, and filtering them to obtain a mixed alcohol extract and mixed medicinal residues, recovering ethanol from the mixed alcohol extract to obtain a mixed clear paste;

[0009] 3) freeze-drying and grinding the raw hawthorn, and slowly adding the ground hawthorn powder to the stirring hydrogen peroxide solution. After the addition, ultrasonic heating is performed at 40-50° C. for 2-3 hours to obtain a hawthorn hydrolyzate;

[0010] 4) adding the epimedium dregs and the mixed dregs to the hawthorn hydrolyzate, adding a complex enzyme, adjusting the pH to 3-4 with HCl solution, stirring and hydrolyzing at 50° C., and inactivating the enzyme in a water bath after the hydrolysis is completed to obtain an enzymatic hydrolyzate; wherein the complex enzyme comprises one or more of pectinase, cellulase, and hemicellulase;

[0011] 5) subjecting the enzymatic hydrolysate to microwave-assisted reflux extraction, filtering, and concentrating the filtrate to obtain a water-extracted clear paste;

[0012] 6) The epimedium paste, the mixed paste and the water-extracted paste are mixed and filtered, gypenosides are added to the filtrate, and water is added to obtain the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction.

[0013] The weight ratio of the raw materials in the composition of the present invention is: 22 parts of ginkgo leaves, 12 parts of salvia miltiorrhiza, 0.02 parts of gypenosides, 1.9 parts of Corydalis yanhusuo, 17 parts of hawthorn, 17 parts of leucanthemum, and 17 parts of epimedium.

[0014] In the preparation method of the present invention, the epimedium is extracted with alcohol alone, preferably with 85% ethanol concentration, which can better realize the extraction of icariin and avoid the problem of poor stability and precipitation when icariin and other glycosides coexist.

[0015] In the production of Chinese medicine oral solutions, in order to improve their clarity, water extraction and alcohol precipitation are generally used to remove water-soluble impurities. During alcohol precipitation, a volume concentration of 65% ethanol solution is usually used to retain the active ingredients in the aqueous solution and remove oligosaccharides and polysaccharides. In step 2) of the present invention, a volume concentration of 65% ethanol solution is used to extract Corydalis yanhusuo, Herba schizonepetae, Ginkgo biloba, and Salvia miltiorrhiza. This not only ensures the dissolution of the active ingredients, but also prevents a large amount of oligosaccharides and polysaccharides from entering the alcohol extraction solution, making it convenient for the polysaccharides to be hydrolyzed into oligosaccharides in subsequent steps and then extracted to increase the content of oligosaccharides in the composition.

[0016] The hawthorn fruit, a raw material of the present invention, is rich in pectin and crude fiber, resulting in high viscosity. This leads to uneven temperature distribution during heating and easily becomes a paste when stirred, forming a viscous gel mass. Direct enzymatic hydrolysis, however, yields unsatisfactory hydrolysis results. The present invention first hydrolyzes the hawthorn fruit with hydrogen peroxide ultrasonically, hydrolyzing the viscous, insoluble pectin and hawthorn polysaccharides into small, soluble hawthorn polysaccharides or hawthorn oligosaccharides with low viscosity. This also promotes the dissolution of active ingredients such as flavonoids, organic acids, and triterpenoid saponins. Furthermore, the vitamin C and organic acids in the hawthorn fruit assist in the degradation of hawthorn pectin by hydrogen peroxide, thereby increasing the efficiency of hawthorn pectin hydrolysis. Therefore, the present invention utilizes a 1-2% by mass hydrogen peroxide solution for ultrasonic hydrolysis of the hawthorn fruit. This reduces the viscosity of the hawthorn hydrolyzate, provides good mixing and stirring, and provides a uniform temperature distribution, thereby increasing the efficiency of subsequent enzymatic hydrolysis. The higher the concentration of the hydrogen peroxide solution, the lower the molecular weight of the resulting hawthorn oligosaccharides. However, when the hydrogen peroxide concentration exceeds 2%, some hawthorn oligosaccharides are hydrolyzed into inactive monosaccharides, which can easily lead to the loss of active antioxidant components in hawthorn. This can also cause residual hydrogen peroxide to affect the activity of the complex enzyme. If the hydrogen peroxide concentration is too low, most of the hydrogen peroxide will be hydrolyzed to form water and oxygen during ultrasonic hydrolysis, failing to achieve a good hydrolysis effect. Therefore, the concentration of the hydrogen peroxide solution should be controlled between 1-2%, with an optimal value of 1.8%.

[0017] Preferably, in step 3), the mass ratio of hydrogen peroxide solution to hawthorn powder is 1:30, the ultrasonic power is 120 W, and the ultrasonic time is 2 h.

[0018] The present invention adds epimedium dregs and mixed dregs to a hawthorn hydrolyzate for composite enzymatic hydrolysis, which can hydrolyze the polysaccharides and pectin in hawthorn, epimedium dregs, and mixed dregs into oligosaccharides and oligosaccharides, and fully hydrolyze and dissolve the polysaccharides in the dregs. The mass ratio of cellulase, pectinase, and hemicellulase in the composite enzyme is 10-20:2-4:5-10; the enzymatic activity of each enzyme in the composite enzyme is 100,000 U / g to 150,000 U / g. The amount of composite enzyme added is 2% of the total weight of the epimedium dregs and mixed dregs, and the stirring and hydrolysis time is 4 hours. Using this ratio of composite enzymes, the hydrolysis effect is good, and the obtained oligosaccharides have a molecular weight of less than 2000 Da, which has good effects on promoting blood circulation, reducing turbidity, removing toxic heavy metal ions, and preventing myocardial infarction.

[0019] Specifically, the hawthorn oligosaccharide has good antioxidant activity and improves blood lipid metabolism. It can reduce the cholesterol and triglyceride levels in the blood and inhibit fat absorption. It also has the effect of inhibiting α-glucosidase activity and increasing β-glucosidase activity. Inhibiting α-glucosidase activity can lower blood sugar concentration and prevent lipid substances in the blood from depositing on the inner walls of blood vessels, thereby avoiding the formation of atherosclerotic plaques. Increasing β-glucosidase activity can promote the conversion of icariin, 2"-O-galloyl-glucosidoside and chloranthus glycosides into absorbable and usable low-sugar glycosides or aglycones, which enter the blood circulation, thereby increasing the bioavailability of icariin, 2"-O-galloyl-glucosidoside and chloranthus glycosides. It also has the effect of lowering total cholesterol and low-density lipoprotein cholesterol levels in plasma, ultimately reducing atherosclerotic plaques and preventing the formation of atherosclerotic plaques. In addition, the carboxyl anions in hawthorn oligosaccharides have a strong ability to bind cations, which can chelate heavy metal ions such as lead, mercury, arsenic, and cadmium, promote the urinary excretion of toxic elements in the blood, thereby avoiding vascular endothelial damage caused by heavy metal ions and ultimately reducing the risk of atherosclerosis and myocardial infarction.

[0020] The ginkgo leaf polysaccharide obtained by enzymatic hydrolysis in the preparation method can inhibit platelet aggregation and thrombosis, improve the phagocytic ability of phagocytes, scavenge free radicals, resist lipid peroxidation, and has anti-thrombotic, immune-enhancing, anti-aging and other effects.

[0021] The salvia miltiorrhiza polysaccharide obtained by enzymatic hydrolysis in this preparation method can increase insulin sensitivity, thereby helping to lower blood sugar, and also has an antioxidant effect. It can also increase coronary blood flow, improve microcirculation, and help alleviate the symptoms of myocardial ischemia.

[0022] In addition, the various oligosaccharides obtained by microwave-assisted reflux extraction of the enzymatic hydrolysate can also increase the number of probiotics in the intestinal flora, inhibit the proliferation of harmful bacteria, significantly proliferate Lactobacillus plantarum, Lactobacillus rhamnosus, Bifidobacterium longum and Bifidobacterium adolescentis, inhibit the growth of Escherichia coli in the intestine, increase the activity and metabolic efficiency of the intestinal flora, and more effectively improve the bioavailability of the active ingredients.

[0023] Therefore, the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction obtained by the above-mentioned preparation method of the present invention has high oligosaccharide and polysaccharide contents. For example, the density of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction obtained in step 6) is 1.01-1.02 g / mL, wherein the total sugar content is 13.46-14.29 mg / mL, the oligosaccharide content is 9.54-9.87 mg / mL, and the polysaccharide content is 3.12-3.58 mg / mL. The composition significantly improves its ability to scavenge blood free radicals, lower blood sugar, and lower plasma low-density lipoprotein cholesterol levels, can comprehensively purify blood, reduce atherosclerotic plaques, and prevent the formation of atherosclerotic plaques.

[0024] In addition, the presence of multiple oligosaccharides in the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction obtained by the above-mentioned preparation method not only solubilizes icariin, 2"-O-galloyl icariin and crystalline orchid, thereby increasing the solubility of the three glycosides, but also improves the storage stability of 2"-O-galloyl icariin and crystalline orchid, thereby avoiding the precipitation of icariin and the degradation of 2"-O-galloyl icariin and crystalline orchid, thereby extending the shelf life of the composition.

[0025] After the present invention adopts such a design, it has at least the following advantages:

[0026] The composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction obtained by the preparation method of the present invention has a high content of oligosaccharides, a high content of glycoside substances, good storage stability, a long shelf life, and high bioavailability. It can significantly reduce the levels of total cholesterol and low-density lipoprotein cholesterol in plasma, reduce atherosclerotic plaques, prevent the formation of atherosclerotic plaques, has good effects of promoting blood circulation, reducing turbidity and preventing myocardial infarction, and also has the technical effect of removing toxic heavy metal ions in the body. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] The above is only an overview of the technical solution of the present invention. In order to more clearly understand the technical means of the present invention, the present invention is further described in detail below in conjunction with the accompanying drawings and specific embodiments.

[0028] Figure 1 This is a bar graph showing the results of the contents of glycosides, oligosaccharides and polysaccharides in the compositions for promoting blood circulation, reducing turbidity and preventing myocardial infarction in Experimental Example 1 of the present invention.

[0029] Figure 2This is a bar graph of the relative areas of atherosclerotic plaques obtained from the compositions for promoting blood circulation, reducing turbidity and preventing myocardial infarction in Experimental Example 3 of the present invention.

[0030] Figure 3 This is a bar graph of the bioavailability of icariin in Experimental Example 4 of the present invention.

[0031] Figure 4 This is a line graph showing changes in total cholesterol concentration in rat plasma for each of the blood-activating, turbidity-reducing and myocardial infarction-preventing compositions in Experimental Example 5 of the present invention.

[0032] Figure 5 This is a line graph showing changes in the concentration of low-density lipoprotein cholesterol in rat plasma for each of the blood-activating, turbidity-reducing and myocardial infarction-preventing compositions in Experimental Example 5 of the present invention.

[0033] Figure 6 This is a line graph showing changes in blood lead concentration in rats with the compositions for promoting blood circulation, reducing turbidity and preventing myocardial infarction in Experimental Example 6 of the present invention. DETAILED DESCRIPTION

[0034] Exemplary embodiments of the present invention will be described in more detail below with reference to the accompanying drawings. Although exemplary embodiments of the present invention are shown in the accompanying drawings, it should be understood that the present invention can be implemented in various forms and should not be limited by the embodiments set forth herein. Rather, these embodiments are provided to enable a more thorough understanding of the present invention and to fully convey the scope of the present invention to those skilled in the art.

[0035] Example 1

[0036] The raw materials of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction are: 220g of Ginkgo biloba leaves, 170g of Herba Epimedii, 170g of Salvia miltiorrhizae, 0.2g of Gynostemma pentaphyllum total glycosides, 19g of Corydalis yanhusuo and 170g of Crataegus pinnatifida. The composition is prepared by the following steps:

[0037] 1) Crush the epimedium, pass it through a pharmacopoeia No. 4 sieve, add 85% ethanol to a solid-to-liquid ratio of 1:25 (w / v), and soak for 20 hours before extraction. The extraction temperature is 60°C, the extraction time is 2 hours, and the extract is filtered to obtain an epimedium alcohol extract and epimedium residue; the ethanol is recovered from the epimedium alcohol extract to obtain an epimedium paste;

[0038] In this example, the conditions for extracting icariin were determined through a single-factor experiment: Epimedium was crushed and passed through a No. 4 sieve. Equal amounts of icariin were then sonicated for 1 hour using 60%, 75%, 80%, 85%, and 90% ethanol solutions by volume. After cooling, the ethanol solution was added to bring the volume back to the mark. The icariin content of each solution was determined using high-performance liquid chromatography. The results showed that the 85% ethanol solution had the highest icariin concentration.

[0039] 2) Crush Corydalis yanhusuo, cyperus rotundus, ginkgo biloba, and salvia miltiorrhiza, pass through a pharmacopoeia No. 4 sieve, add 65% ethanol to a solid-liquid ratio of 1:20 (w / v), soak for 20 hours, and extract at 80°C for 2 hours. Filter to obtain a mixed alcohol extract and mixed medicinal residues, and recover ethanol from the mixed alcohol extract to obtain a mixed clear paste;

[0040] 3) Freeze-dried hawthorn was ground and added with a 1.8% hydrogen peroxide solution to a solid-liquid ratio of 1:30 (w / v). After mixing, the mixture was ultrasonically heated at 120 W of ultrasonic power at 40°C for 2 h to obtain a hawthorn hydrolyzate.

[0041] 4) mixing the epimedium dregs and the mixed dregs and adding them to the hawthorn hydrolyzate, adding 2% of the total weight of the epimedium dregs and the mixed dregs to a complex enzyme, adjusting the pH to 3-4 with HCl solution, the temperature to 50° C., the stirring speed to 135 rpm, and hydrolyzing for 4 h. After the hydrolysis is completed, the enzyme is inactivated in a water bath for 5 min to obtain an enzymatic hydrolyzate;

[0042] The compound enzyme comprises cellulase, pectinase and hemicellulase, and the dosage ratio thereof is 14:2:6; the enzyme activity of each enzyme in the compound enzyme is 150,000 U / g;

[0043] 5) The obtained enzymatic hydrolysate was subjected to microwave-assisted reflux extraction twice, the first reflux extraction was 2 hours, and the second reflux extraction was 1.5 hours, filtered, and the two filtrates were combined and concentrated to a density of 1.02-1.05 g / mL to obtain a water-extracted clear paste;

[0044] 6) The above-obtained epimedium paste, mixed paste and water-extracted paste were mixed and filtered, gypenosides were added to the filtrate, and water was added to a density of 1.01-1.02 g / mL to obtain a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction.

[0045] Example 2

[0046] The raw materials of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction are: 220g of Ginkgo biloba leaves, 170g of Herba Epimedii, 170g of Salvia miltiorrhizae, 0.2g of Gynostemma pentaphyllum total glycosides, 19g of Corydalis yanhusuo and 170g of Crataegus pinnatifida. The composition is prepared by the following steps:

[0047] 1) crushing epimedium, passing through a pharmacopoeia No. 4 sieve, adding 85% ethanol by volume to a solid-liquid ratio of 1:25 (w / v), soaking for 20 hours, and then extracting at 60°C; the extraction time is 2 hours, filtering to obtain an epimedium alcohol extract and epimedium residue; recovering ethanol from the epimedium alcohol extract to obtain an epimedium paste;

[0048] 2) Crush Corydalis yanhusuo, cyperus rotundus, ginkgo biloba, and salvia miltiorrhiza, pass through a pharmacopoeia No. 4 sieve, add 65% ethanol to a solid-liquid ratio of 1:20 (w / v), soak for 20 hours, and extract at 80°C for 2 hours. Filter to obtain a mixed alcohol extract and mixed medicinal residues, and recover ethanol from the mixed alcohol extract to obtain a mixed clear paste;

[0049] 3) Freeze-dried hawthorn was ground and pulverized, and a 1% hydrogen peroxide solution was added to a solid-liquid ratio of 1:30 (w / v). After mixing, the mixture was ultrasonically heated at 50°C for 2 h at 120 W ultrasonic power to obtain a hawthorn hydrolyzate.

[0050] 4) adding the epimedium dregs and the mixed dregs to the hawthorn hydrolyzate, adding 2% of the total weight of the epimedium dregs and the mixed dregs to the complex enzyme, adjusting the pH to 3-4 with HCl solution, the temperature to 50° C., the stirring speed to 135 rpm, and hydrolyzing for 4 h. After the hydrolysis is completed, the enzyme is inactivated in a water bath for 5 min to obtain an enzymatic hydrolyzate;

[0051] The compound enzyme comprises cellulase, pectinase and hemicellulase, and the dosage ratio thereof is 10:3:5; the enzyme activity of each enzyme in the compound enzyme is 100,000 U / g;

[0052] 5) The obtained enzymatic hydrolysate was subjected to microwave-assisted reflux extraction twice, the first reflux extraction was 2 hours, and the second reflux extraction was 1.5 hours, filtered, and the two filtrates were combined and concentrated to a density of 1.02-1.05 g / mL to obtain a water-extracted clear paste;

[0053] 6) The above-obtained epimedium paste, mixed paste and water-extracted paste were mixed and filtered, gypenosides were added to the filtrate, and water was added to a density of 1.01-1.02 g / mL to obtain a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction.

[0054] Example 3

[0055] The raw materials of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction are: 220g of Ginkgo biloba leaves, 170g of Herba Epimedii, 170g of Salvia miltiorrhizae, 0.2g of Gynostemma pentaphyllum total glycosides, 19g of Corydalis yanhusuo and 170g of Crataegus pinnatifida. The composition is prepared by the following steps:

[0056] 1) crushing epimedium, passing through a pharmacopoeia No. 4 sieve, adding 85% ethanol by volume to a solid-liquid ratio of 1:25 (w / v), soaking for 20 hours, and then extracting at 60°C; the extraction time is 2 hours, filtering to obtain an epimedium alcohol extract and epimedium residue; recovering ethanol from the epimedium alcohol extract to obtain an epimedium paste;

[0057] 2) Crush Corydalis yanhusuo, cyperus rotundus, ginkgo biloba, and salvia miltiorrhiza, pass through a pharmacopoeia No. 4 sieve, add 65% ethanol to a solid-liquid ratio of 1:20 (w / v), soak for 20 hours, and extract at 80°C for 2 hours. Filter to obtain a mixed alcohol extract and mixed medicinal residues, and recover ethanol from the mixed alcohol extract to obtain a mixed clear paste;

[0058] 3) Freeze-dried hawthorn was ground and added with a 1.5% hydrogen peroxide solution to a solid-liquid ratio of 1:30 (w / v). After mixing, the mixture was ultrasonically heated at 120 W ultrasonic power and 40°C for 3 h to obtain a hawthorn hydrolyzate.

[0059] 4) adding the epimedium dregs and the mixed dregs to the hawthorn hydrolyzate, adding 2% of the total weight of the epimedium dregs and the mixed dregs to the complex enzyme, adjusting the pH to 3-4 with HCl solution, the temperature to 50° C., the stirring speed to 135 rpm, and hydrolyzing for 4 h. After the hydrolysis is completed, the enzyme is inactivated in a water bath for 5 min to obtain an enzymatic hydrolyzate;

[0060] The compound enzyme comprises cellulase, pectinase and hemicellulase, and the dosage ratio thereof is 20:4:5; the enzyme activity of each enzyme in the compound enzyme is 100,000 U / g;

[0061] 5) The obtained enzymatic hydrolysate was subjected to microwave-assisted reflux extraction twice, the first reflux extraction was 2 hours, and the second reflux extraction was 1.5 hours, filtered, and the two filtrates were combined and concentrated to a density of 1.02-1.05 g / mL to obtain a water-extracted clear paste;

[0062] 6) The above-obtained epimedium paste, mixed paste and water-extracted paste were mixed and filtered, gypenosides were added to the filtrate, and water was added to a density of 1.01-1.02 g / mL to obtain a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction.

[0063] Example 4

[0064] The raw materials of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction are: 220g of Ginkgo biloba leaves, 170g of Herba Epimedii, 170g of Salvia miltiorrhizae, 0.2g of Gynostemma pentaphyllum total glycosides, 19g of Corydalis yanhusuo and 170g of Crataegus pinnatifida. The composition is prepared by the following steps:

[0065] 1) crushing epimedium, passing through a pharmacopoeia No. 4 sieve, adding 85% ethanol by volume to a solid-liquid ratio of 1:25 (w / v), soaking for 20 hours, and then extracting at 60°C; the extraction time is 2 hours, filtering to obtain an epimedium alcohol extract and epimedium residue; recovering ethanol from the epimedium alcohol extract to obtain an epimedium paste;

[0066] 2) Crush Corydalis yanhusuo, cyperus rotundus, ginkgo biloba, and salvia miltiorrhiza, pass through a pharmacopoeia No. 4 sieve, add 65% ethanol to a solid-liquid ratio of 1:20 (w / v), soak for 20 hours, and extract at 80°C for 2 hours. Filter to obtain a mixed alcohol extract and mixed medicinal residues, and recover ethanol from the mixed alcohol extract to obtain a mixed clear paste;

[0067] 3) Freeze-dried hawthorn was ground and pulverized, and a 2% hydrogen peroxide solution was added to a solid-liquid ratio of 1:30 (w / v). After mixing, the mixture was ultrasonically heated at 120 W power and 50°C for 2 h to obtain a hawthorn hydrolyzate.

[0068] 4) adding the epimedium dregs and the mixed dregs to the hawthorn hydrolyzate, adding 2% of the total weight of the epimedium dregs and the mixed dregs to the complex enzyme, adjusting the pH to 3-4 with HCl solution, the temperature to 50° C., the stirring speed to 135 rpm, and hydrolyzing for 4 h. After the hydrolysis is completed, the enzyme is inactivated in a water bath for 5 min to obtain an enzymatic hydrolyzate;

[0069] The compound enzyme comprises cellulase, pectinase and hemicellulase, and the dosage ratio thereof is 10:1:5; the enzyme activity of each enzyme in the compound enzyme is 150,000 U / g;

[0070] 5) The obtained enzymatic hydrolysate was subjected to microwave-assisted reflux extraction twice, the first reflux extraction was 2 hours, and the second reflux extraction was 1.5 hours, filtered, and the two filtrates were combined and concentrated to a density of 1.02-1.05 g / mL to obtain a water-extracted clear paste;

[0071] 6) The above-obtained epimedium paste, mixed paste and water-extracted paste were mixed and filtered, gypenosides were added to the filtrate, and water was added to a density of 1.01-1.02 g / mL to obtain a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction.

[0072] Comparative Example 1

[0073] The raw materials of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction are: 220g of Ginkgo biloba leaves, 170g of Herba Epimedii, 170g of Salvia miltiorrhizae, 0.2g of Gynostemma pentaphyllum total glycosides, 19g of Corydalis yanhusuo and 170g of Crataegus pinnatifida. The composition is prepared by the following steps:

[0074] 1) Crush the epimedium, pass it through a pharmacopoeia No. 4 sieve, add 85% ethanol to a solid-to-liquid ratio of 1:25 (w / v), and soak for 20 hours before extraction. The extraction temperature is 60°C, the extraction time is 2 hours, and the extract is filtered to obtain an epimedium alcohol extract and epimedium residue; the ethanol is recovered from the epimedium alcohol extract to obtain an epimedium paste;

[0075] 2) Crush Corydalis yanhusuo, cyperus rotundus, ginkgo biloba, and salvia miltiorrhiza, pass through a pharmacopoeia No. 4 sieve, add 65% ethanol to a solid-liquid ratio of 1:20 (w / v), soak for 20 hours, and extract at 80°C for 2 hours. Filter to obtain a mixed alcohol extract and mixed medicinal residues, and recover ethanol from the mixed alcohol extract to obtain a mixed clear paste;

[0076] 3) Freeze-dried hawthorn was ground and water was added to a solid-liquid ratio of 1:30 (w / v). Ultrasonic heating was performed at 70°C for 2 h at 120 W ultrasonic power to obtain a hawthorn extract.

[0077] 4) mixing the epimedium dregs and the mixed dregs and adding them to the hawthorn extract, adding 2% of the total weight of the epimedium dregs and the mixed dregs as a complex enzyme, adjusting the pH to 3-4 with HCl solution, the temperature to 50° C., the stirring speed to 135 rpm, and hydrolyzing for 4 h. After the hydrolysis is completed, the enzyme is inactivated in a water bath for 5 min to obtain an enzymatic hydrolyzate;

[0078] The compound enzyme comprises cellulase, pectinase and hemicellulase, and the dosage ratio thereof is 14:2:6; the enzyme activity of each enzyme in the compound enzyme is 150,000 U / g;

[0079] 5) The obtained enzymatic hydrolysate was subjected to microwave-assisted reflux extraction twice, the first reflux extraction was 2 hours, and the second reflux extraction was 1.5 hours, filtered, and the two filtrates were combined and concentrated to a density of 1.02-1.05 g / mL to obtain a water-extracted clear paste;

[0080] 6) The above-obtained epimedium paste, mixed paste and water-extracted paste were mixed and filtered, gypenosides were added to the filtrate, and water was added to a density of 1.01-1.02 g / mL to obtain a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction.

[0081] Comparative Example 2

[0082] The raw materials of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction are: 220g of Ginkgo biloba leaves, 170g of Herba Epimedii, 170g of Salvia miltiorrhizae, 0.2g of Gynostemma pentaphyllum total glycosides, 19g of Corydalis yanhusuo and 170g of Crataegus pinnatifida. The composition is prepared by the following steps:

[0083] 1) Crush the epimedium, pass it through a pharmacopoeia No. 4 sieve, add 85% ethanol to a solid-to-liquid ratio of 1:25 (w / v), and soak for 20 hours before extraction. The extraction temperature is 60°C, the extraction time is 2 hours, and the extract is filtered to obtain an epimedium alcohol extract and epimedium residue; the ethanol is recovered from the epimedium alcohol extract to obtain an epimedium paste;

[0084] 2) Crush Corydalis yanhusuo, cyperus rotundus, ginkgo biloba, and salvia miltiorrhiza, pass through a pharmacopoeia No. 4 sieve, add 65% ethanol to a solid-liquid ratio of 1:20 (w / v), soak for 20 hours, and extract at 80°C for 2 hours. Filter to obtain a mixed alcohol extract and mixed medicinal residues, and recover ethanol from the mixed alcohol extract to obtain a mixed clear paste;

[0085] 3) Freeze-dried hawthorn was ground and added with a 1.8% hydrogen peroxide solution to a solid-liquid ratio of 1:30 (w / v). After mixing, the mixture was ultrasonically heated at 120 W of ultrasonic power at 40°C for 2 h to obtain a hawthorn hydrolyzate.

[0086] 4) mixing the epimedium residue and the mixed residue and adding them to the hawthorn hydrolyzate, heating at 50° C. and stirring at 135 r / min for 4 h to obtain a mixed extract;

[0087] 5) The obtained mixed extract was subjected to microwave-assisted reflux extraction twice, the first reflux extraction was 2 hours, and the second reflux extraction was 1.5 hours, filtered, and the two filtrates were combined and concentrated to a density of 1.02-1.05 g / mL to obtain a water-extracted clear paste;

[0088] 6) The above-obtained epimedium paste, mixed paste and water-extracted paste were mixed and filtered, gypenosides were added to the filtrate, and water was added to a density of 1.01-1.02 g / mL to obtain a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction.

[0089] Comparative Example 3

[0090] The raw materials of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction are: 220g of Ginkgo biloba leaves, 170g of Herba Epimedii, 170g of Salvia miltiorrhizae, 0.2g of Gynostemma pentaphyllum total glycosides, 19g of Corydalis yanhusuo and 170g of Crataegus pinnatifida. The composition is prepared by the following steps:

[0091] 1) Crush the epimedium, pass it through a pharmacopoeia No. 4 sieve, add 85% ethanol to a solid-to-liquid ratio of 1:25 (w / v), and soak for 20 hours before extraction. The extraction temperature is 60°C, the extraction time is 2 hours, and the extract is filtered to obtain an epimedium alcohol extract and epimedium residue; the ethanol is recovered from the epimedium alcohol extract to obtain an epimedium paste;

[0092] 2) Crush Corydalis yanhusuo, cyperus rotundus, ginkgo biloba, and salvia miltiorrhiza, pass through a pharmacopoeia No. 4 sieve, add 65% ethanol to a solid-liquid ratio of 1:20 (w / v), soak for 20 hours, and extract at 80°C for 2 hours. Filter to obtain a mixed alcohol extract and mixed medicinal residues, and recover ethanol from the mixed alcohol extract to obtain a mixed clear paste;

[0093] 3) Freeze-dried hawthorn was ground and water was added to a solid-liquid ratio of 1:30 (w / v). Ultrasonic heating was performed at 70°C for 2 h at 120 W ultrasonic power to obtain a hawthorn extract.

[0094] 4) mixing the epimedium residue and the mixed residue and adding them to the hawthorn extract, heating at 50° C. and stirring at 135 r / min for 4 h to obtain a mixed extract;

[0095] 5) The obtained mixed extract was subjected to microwave-assisted reflux extraction twice, the first reflux extraction was 2 hours, and the second reflux extraction was 1.5 hours, filtered, and the two filtrates were combined and concentrated to a density of 1.02-1.05 g / mL to obtain a water-extracted clear paste;

[0096] 6) The above-obtained epimedium paste, mixed paste and water-extracted paste were mixed and filtered, gypenosides were added to the filtrate, and water was added to a density of 1.01-1.02 g / mL to obtain a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction.

[0097] Comparative Example 4

[0098] The raw materials of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction are: 220g of Ginkgo biloba leaves, 170g of Herba Epimedii, 170g of Salvia miltiorrhizae, 0.2g of Gynostemma pentaphyllum total glycosides, 19g of Corydalis yanhusuo and 170g of Crataegus pinnatifida. The composition is prepared by the following steps:

[0099] 1) Take the raw material of Deer Herb, crush it, add 60% ethanol solution 10 times the weight of Deer Herb, soak it for 30 hours, then heat and reflux to extract once, the extraction time is 1.5 hours, then filter the extract, retain the Deer Herb residue for later use, and concentrate the filtrate under reduced pressure to a Deer Herb paste with a relative density of 1.15 (50°C) for later use.

[0100] 2) Take the raw material Epimedium, crush it, add 65% ethanol solution 10 times the weight of Epimedium, soak it for 30 hours, then heat and reflux extract once, the extraction time is 1.5 hours, then filter the extract, retain the Epimedium residue for later use, and concentrate the filtrate under reduced pressure to an Epimedium paste with a relative density of 1.15-1.20 (50° C.) for later use.

[0101] 3) Crush the raw materials ginkgo leaf, salvia miltiorrhiza, corydalis yanhusuo and hawthorn separately, and mix the ginkgo leaf crushed material, salvia miltiorrhiza crushed material, corydalis yanhusuo crushed material, hawthorn crushed material with the above-obtained deer mouth herb residue and epimedium residue, add 3000mL of water and soak for 30 minutes. Then add water and boil three times: the first time, add water 8 times the weight of the total amount of soaked medicinal materials, boil for 1.5 hours; the second time, add water 5 times the weight of the total amount of soaked medicinal materials, boil for 1 hour; the third time, add water 4 times the weight of the total amount of soaked medicinal materials, boil for 45 minutes, combine the three decoctions, filter, and concentrate the filtrate under reduced pressure to a mixed clear paste with a relative density of 1.25 (60°C), then cool and set aside.

[0102] 4) The mixed paste, the herbaceous antler paste and the epimedium paste were mixed, and ethanol was added to make the ethanol content reach 65 wt %, followed by standing and filtering. The filtrate was used to recover ethanol and concentrated to a thick paste with a relative density of 1.30 (60° C.).

[0103] 5) Dissolve the raw material gypenosides in 10 mL of water and add it to the thick paste to obtain a composition.

[0104] Experimental Example 1 Detection of the content of active ingredients in each composition

[0105] The compositions prepared in Examples 1-4 and Comparative Examples 1-4 were tested for the contents of the active ingredients icariin, 2"-O-galloyl icariin, icariin, oligosaccharides, and polysaccharides by high performance liquid chromatography. The results are shown in Tables 1 and Figure 1 .

[0106] Table 1 Results of the content of glycosides, oligosaccharides and polysaccharides in each composition (mg / mL)

[0107]

[0108] From Table 1 and Figure 1 It can be seen that the contents of icariin, 2"-O-galloyl-glucosidoside, chloranthus glycosides, oligosaccharides and polysaccharides in the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction obtained by the preparation method of Examples 1-4 are higher than those in the compositions obtained in Comparative Examples 1-4, especially the content of oligosaccharides is particularly obvious.

[0109] The results show that when the concentration of oligosaccharides and polysaccharides in the composition is high, it is beneficial to improve the solubility of 2"-O-galloyl-glucosidoside, chloranthus glycoside and icariin, reduce the amount of precipitation of 2"-O-galloyl-glucosidoside, chloranthus glycoside and icariin, and avoid the problem of loss of effective ingredients during filtration, ultimately significantly increasing the content of effective ingredients in the composition.

[0110] Experimental Example 2 Accelerated stability test of each composition

[0111] The compositions obtained in Examples 1-4 of the present invention and Comparative Examples 1-4 were sealed and subpackaged in brown light-proof bottles, 10 ml / bottle. The bottles were placed in a dark place at a temperature of 40±2°C and a relative humidity of 75±5% for 3 months. Samples were taken every other month to test the contents of icariin, 2"-O-galloyl-glucosidoside, and chloranthroside, and to investigate the attenuation of icariin, 2"-O-galloyl-glucosidoside, and chloranthroside in an accelerated stability test. The results are shown in Table 2.

[0112] Table 2 Changes in the contents of icariin, 2"-O-galloyl icariin and icariin in the accelerated stability test

[0113]

[0114] As can be seen from Table 2 above, the icariin in the compositions for promoting blood circulation, reducing turbidity and preventing myocardial infarction in Examples 1-4 of the present invention has good stability and basically no attenuation. 2"-O-galloyl icariin attenuated 0.70%-2.08% at the end of the third month in the accelerated stability test, and crystal orchid attenuated 1.40%-2.49% at the end of the third month in the accelerated stability test, but the attenuation was less than 5%, which met the requirements of the product quality standards and had excellent storage stability. In Comparative Examples 1-4, icariin attenuated 3.31%-4.71% at the end of the third month in the accelerated stability test, 2"-O-galloyl icariin attenuated 9.66%-17.06% at the end of the third month in the accelerated stability test, and crystal orchid attenuated 16.47%-23.47% at the end of the third month in the accelerated stability test.

[0115] The results show that the storage stability of the compositions for promoting blood circulation, reducing turbidity and preventing myocardial infarction obtained in Examples 1-4 of the present invention is greatly improved as the content of oligosaccharides and polysaccharides increases.

[0116] Experimental Example 3 Effects of each composition on rat atherosclerosis model induced by high-fat diet and vitamin D3

[0117] Atherosclerosis is one of the leading causes of myocardial infarction (MI). When atherosclerosis develops in the coronary arteries of the heart, lipid deposits and plaque formation on the vascular lining can cause narrowing or blockage of the vessels, reducing or blocking the blood supply to the myocardium and causing myocardial ischemia. If the blood supply is interrupted, myocardial cells may die, resulting in MI.

[0118] High doses of vitamin D3 can significantly increase blood calcium. High blood calcium can synergize with high blood lipids to undermine the integrity of the arterial endothelium, thereby facilitating the invasion, damage, and deposition of plasma lipids on the arterial wall. Elevated blood calcium can also promote calcium deposition in the aortic media, promoting smooth muscle cell degeneration and calcification. Therefore, this application uses a high-fat diet combined with vitamin D3 to establish a rat model of atherosclerosis.

[0119] After one week of free diet adaptation, the rats were divided into 10 groups, each with 8 rats. One group was a blank control group, which was fed with ordinary feed (by mass fraction, including: 50% corn flour, 20% soybean meal, 10% wheat bran, 5% fish meal, 5% bone meal, 5% skim milk powder, 5% vitamins and minerals) and had free access to water. The other 9 groups were the model control group, Example 1-4 groups, and Comparative Example 1-4 groups, all of which were fed with high-fat feed (by mass fraction, including: 79.5% ordinary feed, 10% lard, 5% cholesterol, 5% white sugar, 0.5% sodium cholate). At the same time, 6×10 5 The mice were fed with 100 IU / kg vitamin D3 and 0.2% propylthiouracil by gavage at the rate of food intake, with free access to water. The feeding lasted for 60 days, at which point the model was established.

[0120] After the modeling was completed, the Example 1-4 group and the Comparative Example 1-4 group were gavaged with the composition of each Example 1-4 and Comparative Example 1-4, respectively, at a gavage volume of 2 mL / time, twice a day. The model control group was gavaged with an equal amount of normal saline, and the high-fat feed was not stopped during the experiment. On the 0th day, 14th day, and 28th day after the start of the experiment, 10 μL of blood was collected from the orbits of the rats in the Example 1-4 group and the Comparative Example 1-4 group, respectively, for the detection of total cholesterol and low-density lipoprotein cholesterol levels in the plasma in the following Example 5. After 30 days, the animals in each group were fasted overnight and anesthetized by intraperitoneal injection of sodium barbital. Blood was collected from the right femoral artery and serum was separated. The aortic arch, the entire thoracic aorta and the abdominal aorta were stripped off, fixed with a neutral formalin calcium solution, stained with Oil Red O, and then photographed and analyzed for the relative area of ​​atherosclerotic plaques. Relative area = total area of ​​atherosclerotic plaques / area of ​​the aortic intima. The results are shown in Table 3 and the attached Figure 2 .

[0121] Table 3 Relative areas of atherosclerotic plaques in rats of each group.

[0122] Group Relative area of ​​atherosclerotic plaques (%) Blank control group 0 Model control group 44.2±9.1 Example 1 group 7.3±1.5 Example 2 group 7.7±1.9 Example 3 group 7.4±2.0 Example 4 Group 7.5±2.2 Comparative Example 1 15.7±5.6 Comparative Example 2 16.2±4.8 Comparative Example 3 17.5±5.3 Comparative Example 4 19.0±3.4

[0123] From Table 3 and Figure 2It can be seen that the relative areas of atherosclerotic plaques in rats obtained from Example 1-4 and Comparative Example 1-4 groups were significantly decreased compared with the model control group. The results show that the compositions obtained from Example 1-4 and Comparative Example 1-4 can effectively inhibit the formation of atherosclerotic plaques and have a good therapeutic effect on atherosclerosis. The experimental results of Comparative Example 1-4 group were compared with those of Example 1-4 group, and there was still a significant difference in the results. The results show that the composition obtained by the preparation method of Example 1-4 has a significant therapeutic effect on inhibiting the formation of atherosclerotic plaques due to the significant increase in the content of polysaccharides and oligosaccharides, as well as the increase in the content of icariin, 2"-O-galloyl icariin and icariin. It also shows that the content of oligosaccharides and polysaccharides does have an important influence on the efficacy of the composition and can significantly improve the efficacy of the composition.

[0124] Experimental Example 4 Bioavailability Experiment

[0125] On the basis of the above-mentioned Experimental Example 3, the 24-hour excrement (feces and urine) of rats from Groups 1-4 of Examples and Groups 1-4 of Comparative Examples were collected on the 22nd day after continuous administration for 21 days. The urine samples were measured in volume and stored in a -20°C refrigerator; the fecal samples were placed in a cool place to dry and then stored in a -20°C refrigerator. The dried fecal samples were added with 20 times the amount of physiological saline and ground to obtain fecal liquid. The fecal liquid and urine were centrifuged to obtain the supernatant, and ethyl acetate was added for extraction. The ethyl acetate was removed by distillation under reduced pressure, and methanol was added for dissolution. After filtration, the mixture was analyzed by high performance liquid chromatography. The bioavailability of the active ingredients in the composition was analyzed using icariin as the detection index. The results are shown in Table 4 and Attachment. Figure 3 .

[0126] Table 4 Effects of different oral administration of equal doses of drugs on the bioavailability of icariin in rats

[0127] project Urine mg / head Feces mg / head Dosage mg / piece Bioavailability% Example 1 group 1.91±0.08 8.46±1.41 12.76 18.73％ Example 2 group 2.05±0.10 8.13±1.85 12.60 19.21％ Example 3 group 1.87±0.11 8.31±1.55 12.64 19.46％ Example 4 Group 1.62±0.09 8.44±1.76 12.48 19.39％ Comparative Example 1 1.26±0.16 8.61±1.51 11.04 10.60％ Comparative Example 2 1.29±0.18 8.89±1.67 11.36 10.39％ Comparative Example 3 1.24±0.14 8.48±1.60 10.88 10.66％ Comparative Example 4 1.23±0.12 6.75±1.67 8.92 10.54％

[0128] As shown in Table 4 above, the bioavailability of icariin in the compositions for promoting blood circulation, reducing turbidity, and preventing myocardial infarction in Examples 1-4 of the present invention is higher than that in Comparative Examples 1-4. This indicates that the polysaccharide and oligosaccharide content in the compositions prepared by the methods of Examples 1-4 of the present invention is positively correlated with the bioavailability of icariin.

[0129] The results showed that as the content of oligosaccharides and polysaccharides in the composition increased, the bioavailability of icariin increased, ultimately enhancing the effect of the composition in promoting blood circulation, reducing turbidity and preventing myocardial infarction.

[0130] Experimental Example 5 Effects of Each Composition on Plasma Total Cholesterol and Low Density Lipoprotein Cholesterol Levels in Rats

[0131] Based on the above experimental example 3, blood was collected from the orbits of rats in the Example 1-4 group and the Comparative Example 1-4 group on the 0th day, the 14th day, and the 28th day after the start of the experiment. Specifically, the plasma was separated and the total cholesterol and low-density lipoprotein cholesterol levels in the rat plasma were determined using existing kits. The results are shown in Table 5 and the attached Figure 4 、 5 .

[0132] Table 5 Results of total cholesterol and low-density lipoprotein cholesterol concentrations in rat plasma

[0133]

[0134] From the above table 5 and Figure 4 、 5 It can be seen that after 28 days of oral administration, the reduction in the total cholesterol content and low-density lipoprotein cholesterol content in the plasma of rats in Example 1-4 groups was greater than that in Comparative Example 1-4 groups. It can also be seen that the effect of Comparative Example 1 and 2 in reducing the total cholesterol content and low-density lipoprotein cholesterol content in rat plasma was better than that in Comparative Example 3 and 4 groups.

[0135] The above results show that the effect of the composition on reducing the total cholesterol and low-density lipoprotein cholesterol levels in rat plasma is positively correlated with the content of oligosaccharides and polysaccharides. The higher the oligosaccharide and polysaccharide content, the better the effect of reducing the total cholesterol and low-density lipoprotein cholesterol levels in rat plasma. The composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction obtained by the preparation method of the present invention has the effect of significantly reducing the total cholesterol and low-density lipoprotein cholesterol levels in plasma.

[0136] Experimental Example 6 Effects of Each Composition on Blood Lead Ion Content in Preventive High Lead Rat Model

[0137] Take healthy adult male rats weighing 150-200g, after one week of free diet adaptability feeding, the rats are randomly divided into 10 groups, 8 in each group, wherein 1 group is a healthy control group, free drinking of purified water. The other 9 groups are respectively a high lead control group, embodiment 5-8 group, and comparative example 5-8 group, all free drinking of 0.2% lead acetate aqueous solution. All 10 groups of rats are fed ordinary feed (by mass fraction, including: 50% corn flour, 20% soybean meal, 10% wheat bran, 5% fish meal, 5% bone meal, 5% skim milk powder, 5% vitamins and minerals). A total of 30 days of feeding are completed to complete the modeling.

[0138] After the modeling was completed, Example 5-8 Group and Comparative Example 5-8 Group were gavaged with the composition of Example 1-4 and Comparative Example 1-4, respectively, at a gavage volume of 2 mL / time, twice a day. The high-lead control group was gavaged with an equal amount of normal saline. During the experiment, the healthy control group drank purified water freely, and the other groups continued to drink 0.2% lead acetate aqueous solution freely. On the 0th day, 15th day, and 30th day after the start of the experiment, 10 μL of blood was collected from the orbits of the rats in Example 5-8 Group and Comparative Example 5-8 Group, respectively. The lead ion concentration in the rat blood was determined using a graphite furnace atomic absorption spectrophotometer, and the results are shown in Table 6 and Appendix. Figure 6 .

[0139] Table 6 Lead ion content in rat blood

[0140]

[0141] From the above table 6 and Figure 6 As can be seen, the lead removal effects of Examples 5-8 were far superior to those of Comparative Examples 5-8. Furthermore, the trends in blood lead ion concentration in Comparative Examples 7 and 8 were similar to those in the high-lead model group, indicating that the polysaccharide and oligosaccharide content significantly impacted the lead removal effect of the composition. Higher polysaccharide and oligosaccharide content resulted in better lead removal effects, while lower polysaccharide and oligosaccharide content resulted in poorer results. This demonstrates that the composition obtained by the preparation method of the present invention has excellent heavy metal ion removal effects.

[0142] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. Those skilled in the art can make some simple modifications, equivalent changes or modifications based on the technical content disclosed above, which all fall within the scope of protection of the present invention.

Claims

1. A method for preparing a composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction, characterized in that: The raw materials of the composition include ginkgo leaves, salvia miltiorrhiza, total gynostemma glycosides, corydalis yanhusuo, hawthorn, herba damias and epimedium, and the weight ratio of the raw materials in the composition is: 22 parts of ginkgo leaves, 12 parts of salvia miltiorrhiza, 0.02 parts of total gynostemma glycosides, 1.9 parts of corydalis yanhusuo, 17 parts of hawthorn, 17 parts of herba damias and 17 parts of epimedium; The preparation method of the composition comprises the following steps: 1) crushing raw material Epimedium, soaking it in an ethanol solution with a concentration greater than 80%, heating and refluxing for extraction, filtering to obtain Epimedium alcohol extract and Epimedium residue; recovering ethanol from the Epimedium alcohol extract to obtain Epimedium clear paste; 2) crushing the raw materials Corydalis yanhusuo, gypsophila gracile, ginkgo biloba, and salvia miltiorrhiza, mixing them uniformly, soaking them in a 60-70% ethanol solution, heating and refluxing them, and filtering them to obtain a mixed alcohol extract and mixed medicinal residues, recovering ethanol from the mixed alcohol extract to obtain a mixed clear paste; 3) freeze-drying and grinding the raw hawthorn fruit, and slowly adding the ground hawthorn powder to a stirring hydrogen peroxide solution. After the addition, ultrasonically heating at 40-50° C. for 2-3 hours to obtain a hawthorn hydrolyzate; wherein the mass concentration of the hydrogen peroxide solution is 1-2%, the mass ratio of the hydrogen peroxide solution to the hawthorn powder is 1:30, and the ultrasonic power is 120 W; 4) adding the epimedium dregs and the mixed dregs to the hawthorn hydrolyzate, adding a complex enzyme, adjusting the pH to 3-4 with HCl solution, stirring and hydrolyzing at 50° C., and inactivating the enzyme in a water bath after the hydrolysis is completed to obtain an enzymatic hydrolyzate; wherein the complex enzyme comprises one or more of pectinase, cellulase, and hemicellulase; 5) subjecting the enzymatic hydrolysate to microwave-assisted reflux extraction, filtering, and concentrating the filtrate to obtain a water-extracted clear paste; 6) The epimedium paste, the mixed paste and the water-extracted paste are mixed and filtered, gypenosides are added to the filtrate, and water is added to obtain the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction.

2. The method for preparing the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction according to claim 1, characterized in that: The ethanol concentration of the epimedium extracted in step 1) is 85%; the ethanol concentration of the yanhusuo, gypsophila gracile, ginkgo biloba leaves and salvia miltiorrhiza extracted in step 2) is 65%.

3. The method for preparing the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction according to claim 2, characterized in that: In the step 4), the amount of the complex enzyme added is 2% of the total weight of the epimedium dregs and the mixed dregs, and the stirring and hydrolysis time is 4 hours.

4. The method for preparing the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction according to claim 3, characterized in that: The mass ratio of cellulase, pectinase and hemicellulase in the complex enzyme is 10-20:2-4:5-10; and the enzyme activity of each enzyme in the complex enzyme is 100,000 U / g-150,000 U / g.

5. The method for preparing the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction according to claim 4, characterized in that: The composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction obtained in step 6) has a density of 1.01-1.02 g / mL, wherein the total sugar content is 13.46-14.29 mg / mL, the oligosaccharide content is 9.54-9.87 mg / mL, and the polysaccharide content is 3.12-3.58 mg / mL.

6. A composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction, characterized in that: The composition is prepared by the preparation method of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction according to any one of claims 1 to 5.

7. Use of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction according to claim 6 in the preparation of a drug for removing toxic heavy metal ions in the body.

8. Use of the composition for promoting blood circulation, reducing turbidity and preventing myocardial infarction according to claim 6 in the preparation of a drug for lowering total cholesterol and low-density lipoprotein cholesterol in plasma.

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