A composition containing panax notoginseng polysaccharide, a preparation method and application thereof

Abstract

The present application provides a kind of containing sanqi polysaccharide composition, preparation method and its application, belong to plant extraction technical field.The present application containing sanqi polysaccharide composition includes: composite polysaccharide extract, salvia miltiorrhiza extract, kudzu extract and red yeast powder.The present application is crushed after sanqi, angelica, rehmannia glutinosa, ginseng, platycodon grandiflorum and hawthorn is prepared composite polysaccharide extract by removing fat, water extraction, enzymolysis, alcohol precipitation, drying and other steps, salvia miltiorrhiza, kudzu and 70-80% ethanol are mixed to extract, and salvia miltiorrhiza and kudzu extract are dried;Composite polysaccharide extract, salvia miltiorrhiza and kudzu extract and red yeast powder are mixed to obtain sanqi polysaccharide composition.The composite polysaccharide extract prepared by the present application has high hypolipidemic activity, can significantly reduce the body weight and liver weight of hyperlipidemic rats, will not inhibit the normal feeding of hyperlipidemic rats, can improve the HDL-C level in serum, reduce the TC content, TG content and LDL-C content in serum.

Description

Technical Field

[0001] This invention belongs to the field of plant extraction technology, and particularly relates to a composition containing Panax notoginseng polysaccharide, its preparation method and its application. Background Technology

[0002] Hyperlipidemia refers to an abnormality in lipid metabolism in the human body, where the concentration of one or more components of lipids in the blood plasma exceeds the upper limit of normal. It is one of the main pathogenic factors of coronary heart disease and atherosclerosis. Blood contains lipids, including cholesterol, triglycerides, and phospholipids, all of which are insoluble in water and must bind to certain specific proteins (apolipoproteins) to form lipoproteins. Common changes in hyperlipidemia are manifested by abnormalities in one or more of the following indicators: 1. Elevated serum total cholesterol (TC) levels; 2. Elevated serum triglyceride (TG) levels; 3. Abnormally decreased serum high-density lipoprotein cholesterol (HDL-C) levels.

[0003] Currently, the main drugs used in clinical treatment of hyperlipidemia are chemically synthesized drugs, but long-term use of these drugs can cause varying degrees of toxic side effects. Therefore, there is an increasing trend towards finding lipid-lowering substances from natural sources. Polysaccharides have attracted attention due to their good biocompatibility and low toxicity. Polysaccharides are a class of biological macromolecules widely found in plant cell walls, animal cell membranes, and microbial cell walls. Numerous studies have shown that naturally derived polysaccharides possess various biological activities, including anti-tumor, immune-regulating, antioxidant, hypoglycemic, and lipid-lowering effects.

[0004] Currently, a large amount of literature focuses on the study of the structure and bioactivity of single polysaccharides. The lipid-lowering activity of single polysaccharides is limited. Therefore, there is an urgent need to develop a method for separating and purifying medicinal and edible compound polysaccharides, so that the prepared compound polysaccharides can be used to lower blood lipids. Moreover, most compound polysaccharide preparations involve first preparing polysaccharide extracts of each component separately and then mixing them. The preparation method is complicated, the operation is troublesome, and the extraction rate is low. Summary of the Invention

[0005] In view of this, the purpose of the present invention is to provide a composition containing Panax notoginseng polysaccharide, a preparation method thereof, and its application, so as to solve the technical problems of large side effects of chemically synthesized drugs, limited lipid-lowering activity of single polysaccharides, and complex extraction methods and low extraction rates of complex polysaccharides.

[0006] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0007] This invention provides a composition containing Panax notoginseng polysaccharides, the composition comprising:

[0008] Complex polysaccharide extract, tanshinone extract, kudzu root extract, and red yeast rice powder;

[0009] The complex polysaccharide extract includes Panax notoginseng polysaccharide extract, Angelica sinensis polysaccharide extract, Rehmannia glutinosa polysaccharide extract, ginseng polysaccharide extract, Platycodon grandiflorus polysaccharide extract and hawthorn polysaccharide extract;

[0010] The mass ratio of the Panax notoginseng polysaccharide extract, Angelica sinensis polysaccharide extract, Rehmannia glutinosa polysaccharide extract, Ginseng polysaccharide extract, Platycodon grandiflorus polysaccharide extract and Crataegus pinnatifida polysaccharide extract is 7-9:3-6:2-5:1-3:2-4:1-3;

[0011] The mass ratio of the compound polysaccharide extract, tanshinone extract, kudzu root extract and red yeast rice powder is 5-10:4-6:3-8:1-3.

[0012] The present invention also provides a method for preparing the composition containing Panax notoginseng polysaccharides, comprising the following steps:

[0013] 1) Dry and pulverize Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus and hawthorn to obtain Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus and hawthorn powder; soak the Panax notoginseng, Angelica sinensis, Rehmannia glutinosa and ginseng powder in alkaline ethanol and then reflux extract at 50-70℃ for 1-3 hours, repeat 2-3 times to obtain extract; mix the extract with ether to remove impurities, then adjust the pH to neutral and filter to obtain filtrate;

[0014] 2) Mix the filtrate, platycodon powder and hawthorn powder, add petroleum ether to remove grease, let stand, filter, and dry the solid to obtain the composite powder;

[0015] 3) The complex powder is mixed with water and extracted by ultrasonication, followed by enzymatic hydrolysis, and the extract is collected by filtration; the extract is concentrated to obtain a concentrated solution, and anhydrous ethanol is added for alcohol precipitation to obtain a precipitate;

[0016] 4) After redissolving the precipitate in water, add a mixture of chloroform and n-butanol and centrifuge at 5000-6000 r / min for 5-10 min. Collect the supernatant, add anhydrous ethanol again for alcohol precipitation, let stand, centrifuge, wash with acetone, and dry to obtain the complex polysaccharide extract.

[0017] 5) Mix and soak the salvia miltiorrhiza, kudzu root and 70-80% ethanol for 8-15 hours, extract 2-5 times, filter to obtain filtrate, and dry to obtain salvia miltiorrhiza extract and kudzu root extract; mix the compound polysaccharide extract, salvia miltiorrhiza extract, kudzu root extract and red yeast rice powder to obtain a composition containing Panax notoginseng polysaccharide.

[0018] Preferably, in step 1), the mass-to-volume ratio of Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng powder, and alkaline ethanol is 7-9 g: 3-6 g: 2-5 g: 1-3 g: 30-40 mL; the alkaline ethanol is a mixed solution of ethanol and NaOH, wherein the mass concentration of NaOH in the mixed solution is 0.3-0.8%; and the mass concentration of ethanol is 65-75%.

[0019] The volume ratio of the extract to ether is 1:1 to 3; adjust the pH to 6.8 to 7.2.

[0020] Preferably, in step 2), the ratio of filtrate, platycodon powder, hawthorn powder, and petroleum ether is 25-35 mL: 2-4 g: 1-3 g: 45-60 mL; and the settling time is 25-30 min.

[0021] Preferably, in step 3), the mixing ratio of the complex powder to water is 1g: 2-4mL; the ultrasonic power is 300-400W, the ultrasonic temperature is 40-60℃, and the ultrasonic time is 20-25min; the ultrasonic extraction is performed 2-5 times.

[0022] Preferably, the enzyme used for enzymatic hydrolysis in step 3) is a mixture of pectinase, cellulase, and protease in a weight ratio of 2-4:1-3:0.8-1.5; the concentrate is 30%-40% of the extract volume; anhydrous ethanol is added to the concentrate until the ethanol volume concentration is 65%-75%; and the alcohol precipitation time is 8-10 hours.

[0023] After alcohol precipitation, the mixture is centrifuged at a speed of 5000–6000 r / min for a duration of 5–10 min.

[0024] Preferably, in step 4), the volume ratio of chloroform to n-butanol in the chloroform-n-butanol mixture is 3-5:1; anhydrous ethanol is added to the supernatant until the ethanol concentration is 65%-75%; and the standing time is 10-12 hours.

[0025] Preferably, the drying temperature in step 4) is 75-85°C, and the drying time is 4-6 hours.

[0026] Preferably, the drying temperature in step 1) is 40-50°C, and the drying is carried out to a constant weight; the mesh size of the pulverized material is 35-45 mesh.

[0027] The present invention also provides the application of the composition containing Panax notoginseng polysaccharide in the preparation of drugs or functional foods with lipid-lowering effects.

[0028] Compared with the prior art, the present invention has the following beneficial effects:

[0029] The compound polysaccharide extract prepared by this invention has high lipid-lowering activity, can significantly reduce the body weight and liver weight of hyperlipidemic rats, does not inhibit the normal feeding of hyperlipidemic rats, can increase the serum HDL-C level, and reduce the serum TC, TG and LDL-C content.

[0030] This invention removes the toxic component coumarin from Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, and ginseng by alkaline ethanol soaking and acidic solution precipitation. A complex polysaccharide extract is then obtained through ultrasonic extraction, enzymatic hydrolysis, alcohol precipitation, and drying. This method avoids the destruction of active ingredients caused by heating and decoction, resulting in high extraction rates and short extraction times. Simultaneously, the optimized component ratios and extraction process ensure synergistic effects, leading to effective regulation of blood lipids and alleviation of lipid accumulation in the liver. Furthermore, the use of food-grade medicinal materials ensures complete and effective absorption by the body, with no side effects and good safety, avoiding complications and side effects associated with long-term use of Western medicine. Attached Figure Description

[0031] Figure 1 The results of H&E staining of liver tissue in each group are as follows (A: blank group; B: model group; C: positive control group; D: compound low-dose group; E: compound medium-dose group; F: compound high-dose group).

[0032] Figure 2 The results of oil red staining of liver tissue in each group are as follows (A: blank group; B: model group; C: positive control group; D: compound low-dose group; E: compound medium-dose group; F: compound high-dose group). Detailed Implementation

[0033] This invention provides a composition containing Panax notoginseng polysaccharides, the composition comprising:

[0034] Complex polysaccharide extract, tanshinone extract, kudzu root extract, and red yeast rice powder;

[0035] The complex polysaccharide extract includes Panax notoginseng polysaccharide extract, Angelica sinensis polysaccharide extract, Rehmannia glutinosa polysaccharide extract, ginseng polysaccharide extract, Platycodon grandiflorus polysaccharide extract and hawthorn polysaccharide extract;

[0036] The preferred mass ratio of the Panax notoginseng polysaccharide extract, Angelica sinensis polysaccharide extract, Rehmannia glutinosa polysaccharide extract, Ginseng polysaccharide extract, Platycodon grandiflorus polysaccharide extract and Crataegus pinnatifida polysaccharide extract is 7-9:3-6:2-5:1-3:2-4:1-3, and more preferably 8:5:4:2:3:2;

[0037] The preferred mass ratio of the compound polysaccharide extract, tanshinone extract, kudzu root extract and red yeast rice powder is 5-10:4-6:3-8:1-3, and more preferably 8:5:6:2.

[0038] The present invention also provides a method for preparing the composition containing Panax notoginseng polysaccharides, comprising the following steps:

[0039] 1) Dry and pulverize Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus and hawthorn to obtain Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus and hawthorn powder; soak the Panax notoginseng, Angelica sinensis, Rehmannia glutinosa and ginseng powder in alkaline ethanol and then reflux extract at 50-70℃ for 1-3 hours, repeat 2-3 times to obtain extract; mix the extract with ether to remove impurities, then adjust the pH to neutral and filter to obtain filtrate;

[0040] 2) Mix the filtrate, platycodon powder and hawthorn powder, add petroleum ether to remove grease, let stand, filter, and dry the solid to obtain the composite powder;

[0041] 3) The complex powder is mixed with water and extracted by ultrasonication, followed by enzymatic hydrolysis, and the extract is collected by filtration; the extract is concentrated to obtain a concentrated solution, and anhydrous ethanol is added for alcohol precipitation to obtain a precipitate;

[0042] 4) After redissolving the precipitate in water, add a mixture of chloroform and n-butanol and centrifuge at 5000-6000 r / min for 5-10 min. Collect the supernatant, add anhydrous ethanol again for alcohol precipitation, let stand, centrifuge, wash with acetone, and dry to obtain the complex polysaccharide extract.

[0043] 5) Mix and soak the salvia miltiorrhiza, kudzu root and 70-80% ethanol for 8-15 hours, extract 2-5 times, filter to obtain filtrate, and dry to obtain salvia miltiorrhiza extract and kudzu root extract; mix the compound polysaccharide extract, salvia miltiorrhiza extract, kudzu root extract and red yeast rice powder to obtain a composition containing Panax notoginseng polysaccharide.

[0044] In this invention, Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus, and hawthorn are dried and pulverized to obtain powders of Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus, and hawthorn. The drying temperature is preferably 40-50℃, more preferably 42-48℃, and the powder is dried to constant weight. The mesh size of the pulverized powder is preferably 35-45 mesh, more preferably 38-44 mesh. The powders of Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, and ginseng are soaked in alkaline ethanol and then refluxed at 50-70℃ for 1-3 hours. This process is repeated 2-3 times to obtain an extract. The extract is mixed with ether to remove impurities, and then the pH is adjusted to neutral before filtration to obtain a filtrate. The preferred mass-to-volume ratio of Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng powder, and alkaline ethanol is 7-9 g: 3-6 g: 2-5 g: 1-3 g: 30-40 mL, more preferably 8 g: 4-5 g: 3-4 g: 2 g: 32-38 mL; the preferred alkaline ethanol is a mixed solution of ethanol and NaOH, wherein the mass concentration of NaOH in the mixed solution is preferably 0.3-0.8%, more preferably 0.4-0.7%; the preferred mass concentration of ethanol is 65-75%, more preferably 68-72%; the preferred reflux extraction temperature is 55-65℃; the preferred reflux extraction time is 1.5-2.5 h; the preferred number of reflux extractions is 3; the preferred volume ratio of extract to ether is 1:1-3, more preferably 1:2; the preferred pH adjustment is using hydrochloric acid solution to adjust the pH to 6.8-7.2, more preferably 6.9-7.1.

[0045] In this invention, the filtrate, Platycodon grandiflorus powder, and hawthorn powder are mixed and degreased with petroleum ether, allowed to stand, filtered, and the solids are dried to obtain a composite powder. The preferred ratio of the filtrate, Platycodon grandiflorus powder, hawthorn powder, and petroleum ether is 25–35 mL: 2–4 g: 1–3 g: 45–60 mL, more preferably 28–32 mL: 3 g: 2 g: 48–58 mL; the preferred standing time is 25–30 min, more preferably 26–28 min; the preferred drying temperature is 40–60 °C, more preferably 42–58 °C; and the preferred drying time is 1–3 h, more preferably 1.5–2.5 h.

[0046] In this invention, the complex powder is mixed with water for ultrasonic extraction, enzymatic hydrolysis, and the extract is collected after filtration. The extract is concentrated to obtain a concentrate, which is then precipitated with anhydrous ethanol to obtain a precipitate. The preferred mixing ratio of the complex powder to water is 1g:2-4mL, more preferably 1g:3mL; the preferred ultrasonic power is 300-400W, more preferably 320-380W; the preferred ultrasonic temperature is 40-60℃, more preferably 45-55℃; the preferred ultrasonic time is 20-25min, more preferably 22-24min; the preferred number of ultrasonic extractions is 2-5 times, more preferably 3-4 times; the enzyme used for enzymatic hydrolysis is preferably a mixture of pectinase, cellulase, and protease in a weight ratio of 2-4:1-3:0.8-1.5. The preferred weight ratio is 2.5–3.5:1.5–2.5:1–1.3; the concentrate is preferably 30%–40% of the extract volume, more preferably 32%–38%; anhydrous ethanol is added to the concentrate until the ethanol volume concentration is 65%–75%, more preferably 68%–72%; the alcohol precipitation time is preferably 8–10 h, more preferably 8.5–9.5 h; after alcohol precipitation, centrifugation is performed, the centrifugation speed is preferably 5000–6000 r / min, more preferably 5200–5800 r / min; the centrifugation time is preferably 5–10 min, more preferably 7–9 min.

[0047] In this invention, the precipitate is redissolved in water, then centrifuged at 5000-6000 r / min for 5-10 min in a mixture of chloroform and n-butanol. The supernatant is collected, and anhydrous ethanol is added again for alcohol precipitation. After standing, the mixture is centrifuged, washed with acetone, and dried to obtain a complex polysaccharide extract. The volume ratio of chloroform to n-butanol in the chloroform-n-butanol mixture is preferably 3-5:1, more preferably 3.5-4.5:1; anhydrous ethanol is added to the supernatant until the ethanol mass concentration is 65%-75%, more preferably 68%-72%; the settling time is preferably 10-12 hours, more preferably 10.5-11.5 hours; the centrifugation speed is preferably 5000-6000 r / min, more preferably 5200-5800 r / min; the centrifugation time is preferably 5-10 minutes, more preferably 7-9 minutes; the drying temperature is preferably 75-85°C, more preferably 78-82°C; the drying time is preferably 4-6 hours, more preferably 4.5-5.5 hours.

[0048] In this invention, *Salvia miltiorrhiza*, *Pueraria lobata*, and 70-80% ethanol are mixed and soaked for 8-15 hours, extracted 2-5 times, filtered to obtain a filtrate, and dried to obtain *Salvia miltiorrhiza* extract and *Pueraria lobata* extract. A composite polysaccharide extract, *Salvia miltiorrhiza* extract, *Pueraria lobata* extract, and red yeast rice powder are mixed to obtain a composition containing *Panax notoginseng* polysaccharide. The preferred mass concentration of ethanol is 72-78%; the preferred mass-to-volume ratio of *Salvia miltiorrhiza*, *Pueraria lobata*, and ethanol is 4-6 g: 3-8 g: 30-40 mL, more preferably 5 g: 4-7 g: 32-38 mL; the preferred extraction time is 9-14 hours; the preferred number of extractions is 3-4 times; the preferred drying temperature is 75-85℃, more preferably 78-82℃; and the preferred drying time is 4-6 hours, more preferably 4.5-5.5 hours.

[0049] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0050] Example 1

[0051] A composition containing Panax notoginseng polysaccharides, said composition comprising the following components: a complex polysaccharide extract, a Salvia miltiorrhiza extract, a Pueraria lobata extract, and red yeast rice powder;

[0052] The compound polysaccharide extract is Panax notoginseng polysaccharide extract, Angelica sinensis polysaccharide extract, Rehmannia glutinosa polysaccharide extract, ginseng polysaccharide extract, Platycodon grandiflorus polysaccharide extract and hawthorn polysaccharide extract;

[0053] The mass ratio of the Panax notoginseng polysaccharide extract, Angelica sinensis polysaccharide extract, Rehmannia glutinosa polysaccharide extract, Ginseng polysaccharide extract, Platycodon grandiflorus polysaccharide extract, and Crataegus pinnatifida polysaccharide extract is 8:5:4:2:3:2;

[0054] The mass ratio of the compound polysaccharide extract, tanshinone extract, kudzu root extract and red yeast rice powder is 8:5:6:2.

[0055] The preparation method of the composition containing Panax notoginseng polysaccharide is as follows: Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus and hawthorn are dried at 45°C to constant weight and pulverized to 40 mesh to obtain Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus and hawthorn powder; the Panax notoginseng, Angelica sinensis, Rehmannia glutinosa and ginseng powder are soaked in a mixed solution of ethanol and NaOH and then refluxed at 60°C for 2 hours. The mass-volume ratio of the Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng powder and the mixed solution of ethanol and NaOH is 8g:5g:4g:2g:35mL. The mass concentration of NaOH in the mixed solution of ethanol and NaOH is 0.5% and the mass concentration of ethanol is 70%. The extraction is repeated 3 times to obtain the extract. The extract is mixed with ether (volume ratio 1:2) to remove impurities, and then the pH is adjusted to 7 and filtered to obtain the filtrate. The filtrate, platycodon powder, and hawthorn powder were mixed and degreased with petroleum ether. The ratio of the filtrate, platycodon powder, hawthorn powder, and petroleum ether was 30 mL: 3 g: 2 g: 55 mL. The mixture was allowed to stand for 28 minutes, then filtered. The solid was dried at 55°C for 2 hours to obtain the composite powder.

[0056] The complex powder was mixed with water (1g:3mL) and extracted three times by ultrasonication (350W, 50℃, 23min). The enzymes used for enzymatic hydrolysis were pectinase, cellulase and protease in a weight ratio of 3:2:1.2. The extract was collected after filtration. The extract was concentrated to obtain a concentrate, which was 35% of the extract volume. Anhydrous ethanol was added to a concentration of 70% for ethanol precipitation for 9h. The precipitate was obtained by centrifugation (5500r / min, 8min). The precipitate was reconstituted with water and then centrifuged (5500r / min, 8min) with a mixture of chloroform and n-butanol (chloroform:n-butanol = 4:1 (V / V)). The precipitate was collected, and anhydrous ethanol was added again to a concentration of 70% for ethanol precipitation. After standing for 11h, the precipitate was centrifuged (5500r / min, 8min), washed with acetone, and dried (80℃, 5h) to obtain the complex polysaccharide extract.

[0057] The ingredients, including Salvia miltiorrhiza, Pueraria lobata, and 75% ethanol, were mixed and soaked for 12 hours. The extraction was repeated three times. The filtrate was filtered and dried (at 80°C for 5 hours) to obtain Salvia miltiorrhiza extract and Pueraria lobata extract. The mass-volume ratio of Salvia miltiorrhiza, Pueraria lobata, and ethanol was 5 g: 6 g: 35 mL. The compound polysaccharide extract, Salvia miltiorrhiza extract, Pueraria lobata extract, and red yeast rice powder were mixed to obtain a composition containing Panax notoginseng polysaccharide.

[0058] Example 2

[0059] A composition containing Panax notoginseng polysaccharides, said composition comprising the following components: a complex polysaccharide extract, a Salvia miltiorrhiza extract, a Pueraria lobata extract, and red yeast rice powder;

[0060] The compound polysaccharide extract is Panax notoginseng polysaccharide extract, Angelica sinensis polysaccharide extract, Rehmannia glutinosa polysaccharide extract, ginseng polysaccharide extract, Platycodon grandiflorus polysaccharide extract and hawthorn polysaccharide extract;

[0061] The mass ratio of the Panax notoginseng polysaccharide extract, Angelica sinensis polysaccharide extract, Rehmannia glutinosa polysaccharide extract, Ginseng polysaccharide extract, Platycodon grandiflorus polysaccharide extract, and Crataegus pinnatifida polysaccharide extract is 7:3:2:1:2:1;

[0062] The mass ratio of the compound polysaccharide extract, tanshinone extract, kudzu root extract and red yeast rice powder is 5:4:3:1.

[0063] The preparation method of the composition containing Panax notoginseng polysaccharide is as follows: Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus, and hawthorn are dried at 40°C to constant weight and pulverized to 35 mesh to obtain powders of Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus, and hawthorn; the powders of Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, and ginseng are soaked in a mixed solution of ethanol and NaOH and then refluxed at 50°C for 3 hours. The mass-volume ratio of the powders of Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, and ginseng to the mixed solution of ethanol and NaOH is 7g:3g:2g:1g:30mL. The mass concentration of NaOH in the mixed solution of ethanol and NaOH is 0.3%, and the mass concentration of ethanol is 75%. The extraction is repeated twice to obtain an extract. The extract is mixed with ether (volume ratio 1:1) to remove impurities, and then the pH is adjusted to 6.8 and filtered to obtain a filtrate. The filtrate, platycodon powder, and hawthorn powder were mixed and degreased with petroleum ether. The ratio of the filtrate, platycodon powder, hawthorn powder, and petroleum ether was 25 mL: 2 g: 1 g: 45 mL. The mixture was allowed to stand for 25 min, filtered, and the solid was dried at 40 °C for 3 h to obtain the composite powder.

[0064] The complex powder was mixed with water (1g:2mL) and extracted twice by sonication (300W, 40℃, 25min). The enzymes used for enzymatic hydrolysis were pectinase, cellulase and protease in a weight ratio of 2:1:0.8. The extract was collected after filtration. The extract was concentrated to obtain a concentrate, which was 30% of the extract volume. Anhydrous ethanol was added to a concentration of 65% for ethanol precipitation for 8 hours. The precipitate was obtained by centrifugation (5000r / min, 5min). The precipitate was reconstituted with water and then centrifuged (5500r / min, 8min) with a mixture of chloroform and n-butanol (chloroform:n-butanol = 3:1 (V / V)). The precipitate was collected and anhydrous ethanol was added again to a concentration of 65% for ethanol precipitation. After standing for 10 hours, the precipitate was centrifuged (5000r / min, 5min), washed with acetone, and dried (75℃, 4h) to obtain the complex polysaccharide extract.

[0065] The ingredients, including Salvia miltiorrhiza, Pueraria lobata, and 70% ethanol, were mixed and soaked for 8 hours. The extraction was repeated twice. The filtrate was filtered and dried (at 75°C for 4 hours) to obtain Salvia miltiorrhiza extract and Pueraria lobata extract. The mass-volume ratio of Salvia miltiorrhiza, Pueraria lobata, and ethanol was 4g:3g:30mL. The compound polysaccharide extract, Salvia miltiorrhiza extract, Pueraria lobata extract, and red yeast rice powder were mixed to obtain a composition containing Panax notoginseng polysaccharide.

[0066] Example 3

[0067] A composition containing Panax notoginseng polysaccharides, said composition comprising the following components: a complex polysaccharide extract, a Salvia miltiorrhiza extract, a Pueraria lobata extract, and red yeast rice powder;

[0068] The compound polysaccharide extract is Panax notoginseng polysaccharide extract, Angelica sinensis polysaccharide extract, Rehmannia glutinosa polysaccharide extract, ginseng polysaccharide extract, Platycodon grandiflorus polysaccharide extract and hawthorn polysaccharide extract;

[0069] The mass ratio of the Panax notoginseng polysaccharide extract, Angelica sinensis polysaccharide extract, Rehmannia glutinosa polysaccharide extract, Ginseng polysaccharide extract, Platycodon grandiflorus polysaccharide extract, and Crataegus pinnatifida polysaccharide extract is 9:6:5:3:4:3;

[0070] The mass ratio of the compound polysaccharide extract, tanshinone extract, kudzu root extract and red yeast rice powder is 10:6:8:3.

[0071] The preparation method of the composition containing Panax notoginseng polysaccharide is as follows: Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus, and hawthorn are dried at 50°C to constant weight and pulverized to 45 mesh to obtain powders of Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, ginseng, Platycodon grandiflorus, and hawthorn; the powders of Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, and ginseng are soaked in a mixed solution of ethanol and NaOH and then refluxed at 70°C for 1 hour. The mass-volume ratio of the powders of Panax notoginseng, Angelica sinensis, Rehmannia glutinosa, and ginseng to the mixed solution of ethanol and NaOH is 9g:6g:5g:3g:40mL. The mass concentration of NaOH in the mixed solution of ethanol and NaOH is 0.8%, and the mass concentration of ethanol is 65%. The extraction is repeated 3 times to obtain an extract. The extract is mixed with ether (volume ratio 1:3) to remove impurities, and then the pH is adjusted to 7.2 and filtered to obtain a filtrate. The filtrate, platycodon powder, and hawthorn powder were mixed and degreased with petroleum ether. The ratio of the filtrate, platycodon powder, hawthorn powder, and petroleum ether was 35 mL: 4 g: 3 g: 60 mL. The mixture was allowed to stand for 30 min, filtered, and the solid was dried at 60 °C for 1 h to obtain the composite powder.

[0072] The complex powder was mixed with water (1g:4mL) and extracted five times by ultrasonication (400W, 60℃, 20min). The enzymes used for enzymatic hydrolysis were pectinase, cellulase, and protease in a weight ratio of 4:3:1.5. The extract was collected after filtration. The extract was concentrated to obtain a concentrate, which was 40% of the extract volume. Anhydrous ethanol was added to a concentration of 75% for ethanol precipitation for 10h. The precipitate was obtained by centrifugation (6000r / min, 10min). The precipitate was reconstituted with water and then centrifuged (6000r / min, 10min) with a mixture of chloroform and n-butanol (chloroform:n-butanol = 5:1 (V / V)). The precipitate was collected, and anhydrous ethanol was added again to a concentration of 75% for ethanol precipitation. After standing for 12h, the precipitate was centrifuged (5000r / min, 10min), washed with acetone, and dried (85℃, 6h) to obtain the complex polysaccharide extract.

[0073] The ingredients, including Salvia miltiorrhiza, Pueraria lobata, and 80% ethanol, were mixed and soaked for 15 hours. The extraction was repeated 5 times. The filtrate was filtered and dried (at 85°C for 6 hours) to obtain Salvia miltiorrhiza extract and Pueraria lobata extract. The mass-volume ratio of Salvia miltiorrhiza, Pueraria lobata, and ethanol was 6g:8g:40mL. The compound polysaccharide extract, Salvia miltiorrhiza extract, Pueraria lobata extract, and red yeast rice powder were mixed to obtain a composition containing Panax notoginseng polysaccharide.

[0074] Experimental Example 1

[0075] Pitavastatin calcium tablets (Nanjing Changao Pharmaceutical Co., Ltd., batch number 220302, prepared as a 0.1 mg / mL solution in distilled water, prepared every three days); non-assay lipid control product (Shanghai Huizhong Biotechnology Co., Ltd., batch number 201224); cholesterol (TC) assay kit (Shanghai Huizhong Biotechnology Co., Ltd., batch number 01CHOL210226); triglyceride (TG) assay kit (Shanghai Huizhong Biotechnology Co., Ltd., batch number 02TG210330); high-density lipoprotein cholesterol (HDL-c) assay kit (Shanghai Huizhong Biotechnology Co., Ltd., batch number 01HDL210721); low-density lipoprotein cholesterol (LDL-c) assay kit (Shanghai Huizhong Biotechnology Co., Ltd., batch number 01LDL210525).

[0076] SPF-grade male SD rats (Zhejiang Vital River Laboratory Animal Technology Co., Ltd., body weight 180-200g); Ethics Code: PZSHUTCM220711033; housed at the Animal Experiment Center of Shanghai University of Traditional Chinese Medicine, in an environment of 22-26℃ and 40%-60% relative humidity, with free access to food and water. High-fat diet (Wuxi Fanbo Biotechnology Co., Ltd.); maintenance diet (provided by the Animal Experiment Center of Shanghai University of Traditional Chinese Medicine).

[0077] All rats were randomly divided into four groups of 10 each: a blank control group, a high-fat model group, a positive control group (pitavastatin 1 mg / kg), a low-dose combination group (the composition containing Panax notoginseng polysaccharides prepared in Example 1, 50 mg / kg), a medium-dose combination group (the composition containing Panax notoginseng polysaccharides prepared in Example 1, 100 mg / kg), and a high-dose combination group (the composition containing Panax notoginseng polysaccharides prepared in Example 1, 200 mg / kg). The experiment began one week after the rats acclimatized to the environment. The blank control group was fed a maintenance diet, while the other groups were fed a high-fat diet. After 30 days, the rats' body weight was used to assess the model establishment. The rats were then fed according to the experimental design, continuing the feeding process by gavage for another 30 days. At the end of the experiment, rat serum and liver samples were collected for the detection of lipid-lowering related indicators, including body weight, feed intake weight, liver weight, triglycerides (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C).

[0078] 1. Effect of a composition containing Panax notoginseng polysaccharides on the body weight of a high-fat model rat:

[0079] Experimental results are shown in Table 1.

[0080] Table 1. Average body weight of rats in different treatment groups

[0081]

[0082]

[0083] 2. Effects of a composition containing Panax notoginseng polysaccharides on the food intake of a high-fat diet model rat:

[0084] Table 2 Food intake of rats in different treatment groups

[0085]

[0086] As shown in Table 2, the average food intake of rats after modeling was significantly lower than that before modeling. After feeding high-fat model rats with different doses of the composition containing Panax notoginseng polysaccharide prepared in Example 1, the food intake of rats did not change significantly, indicating that the composition of the present invention does not inhibit the normal feeding of rats.

[0087] 3. Effects of a composition containing Panax notoginseng polysaccharides on the organs of a high-fat model rat:

[0088] (1) At the end of the experiment, after the animals were fasted for 10 hours but allowed free access to water, they were anesthetized by intraperitoneal injection of 10% chloral hydrate at 0.35 mL / 100 g body weight. After anesthesia, 5 mL of blood was collected from the abdominal aorta and placed in a 10 mL EP tube. After standing at 4℃ for 4 hours, the tube was centrifuged at 5000 rpm for 10 min, and the supernatant was collected, aliquoted into new 1.5 mL EP tubes, and stored at -80℃ for later use. The liver and epididymal fat were separated, weighed, and used for organ index statistics. The formula for calculating the organ index is: Organ Index (g / g) = Rat organ weight (g) / Rat body weight (g).

[0089] Table 3. Organ indices of high-fat model rats in different treatment groups

[0090]

[0091] As shown in Table 3, feeding high-fat model rats with different doses of the composition containing Panax notoginseng polysaccharide prepared in Example 1 significantly reduced the liver index and epididymal fat index of the rats.

[0092] (2) H&E staining of liver tissue

[0093] After denaturation and coagulation of proteins in liver tissue with 4% paraformaldehyde, the tissue was trimmed and smoothed. The tissue was placed in a dehydration chamber, dehydrated until transparent, and then embedded in paraffin. The embedded paraffin slides were fixed on a microtome and cut into thin sections of approximately 4 μm. The sections were dried in an oven and stored at room temperature for later use. The sections were dewaxed by passing them through xylene and anhydrous ethanol of different concentrations, then stained in hematoxylin solution for about 5 minutes, followed by differentiation with acid and development with ammonia for a few seconds. Then, they were stained with eosin for 1-3 minutes. Finally, the sections were dehydrated in ethanol and xylene until transparent, and resin was dropped onto the transparent sections, which were then sealed with coverslips. Finally, the prepared slides were photographed at 200x magnification.

[0094] Experimental results: such as Figure 1 As shown. By Figure 1 It was found that the livers of the model group rats showed obvious vacuolar degeneration and fatty degeneration of liver tissue cells, and the cell nuclei had deviated from the center of the cells, exhibiting symptoms of moderate fatty liver. Feeding high-fat model rats different doses of the composition containing Panax notoginseng polysaccharide prepared in Example 1 improved the above-mentioned lesions, and the effect was better than that of pitavastatin. This indicates that the composition of the present invention can alleviate fatty degeneration of liver tissue in model animals caused by a high-fat diet to a certain extent.

[0095] (3) Oil Red staining of liver tissue

[0096] After trimming and smoothing the fixed liver tissue, it was dehydrated in 15% and 30% sucrose solutions, respectively. OCT embedding medium was then placed around the completely dehydrated liver tissue for rapid freezing and embedding. The embedded OCT was fixed on a microtome, and sections were cut to a thickness of 8-10 μm. These sections were then mounted on glass slides and stored at -20°C for later use. The frozen sections were dried at room temperature for 10 min; then fixed in paraformaldehyde for 15 min, washed three times with PBS, and incubated in Oil Red working solution in the dark for 10-15 min. After membrane disruption for 10-15 min, the sections were stained again for 1-2 h; stained with 75% ethanol for 2 s, and washed with water for 1 min; counterstained with hematoxylin and differentiated with hydrochloric acid-ethanol, then washed with water. After rinsing, the surrounding moisture was blotted with paper towels, and the sections were mounted with glycerol-gelatin and photographed at 200x magnification.

[0097] Experimental results: such as Figure 2 As shown. By Figure 2 It can be seen that feeding high-fat model rats with different doses of the composition containing Panax notoginseng polysaccharide prepared in Example 1 improved lipid accumulation in the rat liver. The medium dose group showed the most significant reversal of this phenomenon. Pitavastatin was not as effective as the composition containing Panax notoginseng polysaccharide of this invention in treating lipid accumulation in the high-fat model.

[0098] 4. Serum physiological index measurement:

[0099] After 30 days of drug administration and a 16-hour fast, 1 mL of blood was drawn from the eye. The blood was wiped and pressure applied to stop the bleeding before the rats were returned to their cages for continued feeding. Whole blood was left at room temperature for 4 hours, then centrifuged at 5000 rpm at 4°C. The supernatant was aliquoted into 1.5 mL EP tubes. Following strict adherence to the instruction manual and standard operating procedures (SOPs) for the equipment, changes in serum TG, TC, LDL-c, and HDL-c levels in each group of rats were detected using a fully automated biochemical analyzer.

[0100] Experimental results are shown in Table 4.

[0101] Table 4 Serum lipid levels in rats under different treatment groups

[0102] Group TC (mmol / L) TG (mmol / L) HDL-C (mmol / L) LDL-C (mmol / L) Blank group 4.88 0.98 1.13 0.18 Model group 5.42 2.03 0.89 0.39 Positive control group 4.89 1.01 1.21 0.28 Compound low dose 4.97 1.23 1.36 0.31 Compound medium dose 4.85 1.10 1.40 0.26 Compound high dose 4.78 0.92 1.44 0.21

[0103] Table 4 shows that the serum TC, TG, and LDL- levels in rats in the positive drug control group, the low-dose combined group, the medium-dose combined group, and the high-dose combined group were significantly lower than those in the high-fat diet model, while the HDL-C level was significantly higher than that in the high-fat diet model group. With increasing dosage of the composition containing Panax notoginseng polysaccharides, the composition showed better effects in reducing serum TC, TG, and LDL-C levels, even surpassing pitavastatin.

[0104] As can be seen from the above examples and experimental cases, the composition containing Panax notoginseng polysaccharide prepared by the present invention can reduce the body weight of high-fat model rats, reduce the serum TC, TG and LDL- levels of rats and increase HDL-C levels without affecting the normal feeding of rats, and can alleviate the fatty degeneration of liver tissue in model animals caused by high-fat diet, thus having a good lipid-lowering effect.

[0105] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A composition containing Panax notoginseng polysaccharide for lowering blood lipid and relieving liver steatosis, characterized in that, The composition is made of the complex polysaccharide extract, the salvia miltiorrhiza extract, the pueraria extract and the monascus powder; The complex polysaccharide extract is made of the panax notoginseng polysaccharide extract, the angelica polysaccharide extract, the rehmannia polysaccharide extract, the ginseng polysaccharide extract, the platycodon polysaccharide extract and the hawthorn polysaccharide extract; The preparation method of the complex polysaccharide extract comprises the following steps: The panax notoginseng, the angelica, the rehmannia and the ginseng are extracted by refluxing with alkaline ethanol to obtain filtrate; the filtrate, the platycodon and the hawthorn are mixed with petroleum ether to remove fat, and then are extracted by filtration and drying to obtain a complex powder; the complex powder is mixed with water to be extracted by ultrasonic, and then is enzymatically hydrolyzed, concentrated, alcohol precipitated, redissolved with water, added with chloroform-n-butyl alcohol mixture, alcohol precipitated again, washed with acetone and dried to obtain the complex polysaccharide extract; The mass-volume ratio of the panax notoginseng, the angelica, the rehmannia, the ginseng and the alkaline ethanol is 7-9g:3-6g:2-5g:1-3g:30-40mL; The ratio of the filtrate, the platycodon powder and the hawthorn powder to the petroleum ether is 25-35mL:2-4g:1-3g:45-60mL; The mixing ratio of the complex powder to water is 1g:2-4mL; The enzyme used for the enzymatic hydrolysis is a mixture of pectinase, cellulase and protease with a weight ratio of 2-4:1-3:0.8-1.5; The mass ratio of the complex polysaccharide extract, the salvia miltiorrhiza extract, the pueraria extract and the monascus powder is 5-10:4-6:3-8:1-3.

2. The method for preparing the composition containing the panax polysaccharide for reducing blood lipids and relieving liver steatosis according to claim 1, characterized in that, The preparation method comprises the following steps: 1) The panax notoginseng, the angelica, the rehmannia, the ginseng, the platycodon and the hawthorn are dried and crushed to obtain the panax notoginseng, the angelica, the rehmannia, the ginseng, the platycodon and the hawthorn powder; the panax notoginseng, the angelica, the rehmannia and the ginseng powder are soaked in alkaline ethanol and extracted by refluxing at 50-70℃ for 1-3h, and the extraction is repeated for 2-3 times to obtain extract; the extract is mixed with diethyl ether to remove impurities, and then the pH is adjusted to neutral to obtain filtrate; 2) The filtrate, the platycodon powder and the hawthorn powder are mixed with petroleum ether to remove fat, and then are extracted by filtration and drying to obtain a complex powder; 3) The complex powder is mixed with water to be extracted by ultrasonic, and then is enzymatically hydrolyzed, extracted by filtration and collected to obtain extract; the extract is concentrated to obtain concentrated solution, and then is alcohol precipitated with anhydrous ethanol to obtain precipitate; 4) The precipitate is redissolved with water, centrifuged at 5000-6000r / min for 5-10min with chloroform-n-butyl alcohol mixture to obtain supernatant, and then is alcohol precipitated with anhydrous ethanol, centrifuged, washed with acetone and dried to obtain the complex polysaccharide extract; 5) The salvia miltiorrhiza and the pueraria are mixed with 70-80% ethanol to be soaked and extracted for 8-15h, and the extraction is repeated for 2-5 times to obtain filtrate, and then the salvia miltiorrhiza extract and the pueraria extract are obtained by drying; the complex polysaccharide extract, the salvia miltiorrhiza extract, the pueraria extract and the monascus powder are mixed to obtain the composition containing panax notoginseng polysaccharide.

3. The method of claim 2, wherein the preparation of the composition containing the panax polysaccharide for reducing blood lipids and alleviating liver steatosis is characterized by, In step 1), the alkaline ethanol is a mixed solution of ethanol and NaOH, the mass concentration of NaOH in the mixed solution is 0.3-0.8%, and the mass concentration of ethanol is 65-75%; The volume ratio of the extract to diethyl ether is 1:1-3, and the pH is adjusted to 6.8-7.

2.

4. The method of claim 2, wherein the preparation method of the composition containing the panax polysaccharide for reducing blood lipids and alleviating liver steatosis is characterized by, The standing time in step 2) is 25-30 min.

5. The method of claim 2, wherein the preparation method of the composition containing the panax polysaccharide for reducing blood lipids and alleviating liver steatosis is characterized by, The power of the ultrasonic in step 3) is 300-400 W, the temperature of the ultrasonic is 40-60 ℃, and the time of the ultrasonic is 20-25 min; the number of times of the ultrasonic extraction is 2-5.

6. The method of claim 2, wherein the preparation method of the composition containing the panax polysaccharide for reducing blood lipids and alleviating liver steatosis is characterized by, The concentrated liquid in step 3) is 30%-40% of the volume of the extraction liquid; anhydrous ethanol is added to the concentrated liquid to a volume concentration of 65%-75% of ethanol; and the time of the alcohol precipitation is 8-10 h.

7. The method of claim 6, wherein the preparation of the composition containing the panax polysaccharide for reducing blood lipids and alleviating liver steatosis is characterized by, Centrifugation is performed after the alcohol precipitation, the centrifugation speed is 5000-6000 r / min, and the centrifugation time is 5-10 min.

8. The method of claim 2, wherein the preparation method of the composition containing the panax polysaccharide for reducing blood lipids and alleviating liver steatosis is characterized by, The volume ratio of chloroform to n-butanol in the chloroform-n-butanol mixture in step 4) is 3-5:1; anhydrous ethanol is added to the supernatant to a mass concentration of 65%-75% of ethanol; and the standing time is 10-12 h.

9. The method of claim 2, wherein the preparation method of the composition containing the ginseng polysaccharide for reducing blood lipids and alleviating liver steatosis is characterized by, The drying temperature in step 4) is 75-85 ℃, and the drying time is 4-6 h.

10. The method of claim 2, wherein the preparation of the composition containing the ginseng polysaccharide for reducing blood lipids and alleviating liver steatosis is characterized by, The drying temperature in step 1) is 40-50 ℃, and the drying is performed until the weight is constant; and the mesh number of the crushing is 35-45.

11. The use of the composition containing panax polysaccharide for lowering blood lipid and relieving liver steatosis according to claim 1 in the preparation of a medicine for lowering blood lipid and relieving liver steatosis.

Images