WO2022262372A1

WO2022262372A1 - Health-care composition for increasing hdl-c

Info

Publication number: WO2022262372A1
Application number: PCT/CN2022/084858
Authority: WO
Inventors: 林穗珍; 叶锦添; 陈玉嫔; 陈婕思; 黄春晖
Original assignee: 广州市赛普特医药科技股份有限公司
Priority date: 2021-06-16
Filing date: 2022-04-01
Publication date: 2022-12-22
Also published as: CN113350453A; CN113350453B

Abstract

A health-care composition for increasing HDL-C. The composition comprises the following components in percentages by weight: 30-46% of red yeast rice; 10-28% of Ginkgo biloba leaf extract; 0.0005-0.056% of folic acid; 0.001-0.2% of vitamin B6; and 40-50% of an excipient. Compared with the composition of red yeast rice and Ginkgo biloba leaf extract, the health-care composition which has folic acid and vitamin B6 further added thereto can not only significantly reduce the levels of TG, TC and LDL-C in serum, but can also significantly improve the level of HDL-C, so that the health-care composition can more effectively regulate blood fat and be used for preventing various cardiovascular diseases caused by dyslipidemia.

Description

Health care composition for raising HDL-C

technical field

This application relates to a composition suitable for subjects with dyslipidemia, especially a health care composition with the effect of regulating blood lipid, especially raising HDL-C, its preparation method and application.

Background technique

In the past 30 years, the blood lipid level of the Chinese population has gradually increased, and the prevalence of dyslipidemia has increased significantly. According to the 2012 national survey results, the average adult serum total cholesterol (total cholesterol, TC) is 4.50mmol/L; the prevalence of hypercholesterolemia is 4.9%; the average triglyceride (triglyceride, TG) is 1.38mmol/L; The prevalence of hypertriglyceridemia was 13.1%; the average high-density lipoprotein cholesterol (HDL-C) was 1.19mmol/L, and the prevalence of low HDL-C was 33.9%. The overall prevalence of dyslipidemia in Chinese adults is as high as 40.40%, which is a sharp increase compared with 2002. The increase of serum cholesterol level in the population will lead to an increase of about 9.2 million cardiovascular events in my country during 2010-2030.

In 1979, Endo et al. first isolated an active substance called Monacolin K (Monacolin K) from the culture solution of Monascus, which has a strong inhibitory effect on cholesterol synthesis. Subsequent studies have shown that the structure and chemical properties of monacolin K are exactly the same as lovastatin and belong to the same substance. Lovastatin has two forms of acid form and lactone form under acidic conditions, of which the acid form of lovastatin is the pharmacologically active form, because of the ring-opened hydroxy acid part and 3-hydroxy-3-methylpentanediox in its chemical structure Acid monoacyl-CoA has a very similar chemical structure and can competitively inhibit the rate-limiting enzyme of cholesterol biosynthesis—HMG-CoA reductase. Therefore, red yeast rice can reduce cholesterol synthesis, hinder the synthesis and release of very low-density lipoprotein (VLDL) to extrahepatic tissues. On the other hand, monacolin K can compensatory increase the number, activity and affinity of low-density lipoprotein (LDL) receptors on the liver cell membrane through the self-regulating mechanism of liver cells, so that more LDL is metabolized by the LDL receptor pathway, and finally converts cholesterol into bile acid to be excreted, further reducing plasma low-density lipoprotein cholesterol (LDL-C), very low-density lipoprotein cholesterol (VLDL-C) and total cholesterol (TC) s level.

Xuezhikang is a lipid-lowering red yeast rice Chinese patent medicine listed by Beijing Beida Weixin Biotechnology Co., Ltd. According to reports, Xuezhikang is refined from special red yeast rice. The main difference between this red yeast rice and ordinary edible red yeast rice is that it contains a large amount of Monacolin K and its lactone lovastatin. The red yeast rice used in Xuezhikang is fermented from a high-yielding Monacolin-like strain of Monacolin, using japonica rice as raw material, and fermented under special technological conditions, in which the total content of lovastatin reaches more than 20mg/g. The products on the market are divided into capsules and tablets. Each capsule contains 0.3g, orally, 2 capsules at a time, twice a day; each tablet weighs 0.4g, orally, 2 tablets at a time, twice a day.

Different from medicines, the state stipulates that the daily dose of lovastatin in red yeast rice used in health products must be less than 10.0mg/day. Due to the reduced dose of effective active ingredients, the lipid-lowering and lipid-lowering effects of health products are far inferior to those of prescription drugs such as Xuezhikang. In view of this, the prior art provides a composition of red yeast rice and ginkgo biloba extract (Ginko Biloba Extract, GBE) (such as Yinqu capsules, wherein ginkgo biloba extract:red yeast extract=4:1) as a health care product in regulating blood lipids. Studies such as Zhang Chaobo have shown that compared with the model group, Yinqu Capsules can significantly reduce the levels of TC, TG, LDL-C, and TC/HDL-C in rats with hyperlipidemia. g/kg) can increase HDL-C in serum, while HDL-C in the low-dose group (0.5g/kg) has no significant difference (Zhang Chaobo et al., "Studies on the Pharmacodynamics of Yinqu Capsules Regulating Blood Lipids." Anhui Medicine 15.004 ( 2011): 421-423.). It is worth noting that in this study, as a control drug, although the Xuezhikang group (0.2g/kg) had significant differences compared with the model group in terms of reducing TG, TC, and LDL-C, it did not increase HDL -C level.

HDL-C is mainly synthesized in the liver and is an anti-atherosclerotic lipoprotein. HDL transports cholesterol from surrounding tissues (including atherosclerotic plaque) to the liver for recycling or excretion as bile acids, a process known as reverse cholesterol transport. The plasma level of HDL-C is negatively correlated with the risk of atherosclerotic cardiovascular disease (ASCVD). Decreased HDL-C is common in cerebrovascular coronary heart disease, hypertriglyceridemia, liver function damage such as acute and chronic hepatitis, cirrhosis, liver cancer, diabetes, smoking, lack of exercise, etc. Its reduction can be used as a risk indicator for coronary heart disease.

The clinical adverse reactions of GBE involve various systems of the body, such as bleeding (severe intracranial hemorrhage, severe subarachnoid hemorrhage, spontaneous hyphema, severe bilateral subdural hematoma), allergic reaction, hypertension, neutropenia, vascular Redness, seizures, central nervous system excitement, and acute gastroenteritis. The National Institutes of Health (NIH) database shows that the minimum dose of GBE as a dietary supplement per person is 2.5 mg/d, and the maximum dose is 660 mg/d. According to the database of my country's former State Drug and Food Administration, the maximum dose of GBE as a drug is 240mg/d, and the maximum dose as a health food is 200mg/d. In view of the dosage regulations of GBE in health food in the existing regulations, health products obtained by combining GBE with red yeast rice cannot effectively increase HDL-C levels, thus limiting the performance and application range of health products.

Contents of the invention

In view of this, the application provides a health care composition for regulating blood lipids, the dosage of each component of which is in line with the national dosage regulations for each component in health food, and can reduce the levels of TG, TC, LDL-C and Increase the level of HDL-C, so that blood lipids can be more effectively regulated, and it can be used to prevent various cardiovascular diseases caused by abnormal blood lipids.

The inventors of the present application unexpectedly found that adding folic acid and vitamin B6 to the composition of red yeast rice and ginkgo leaf extract can significantly increase the HDL-C level at a low dose of ginkgo leaf extract, while significantly reducing TG, TC, LDL-C levels.

Therefore, one aspect of the present application provides a health care composition for regulating blood lipids, comprising: 30-46% of red yeast rice; 10-28% of ginkgo biloba extract; 0.0005-0.056% of folic acid; vitamin B6 0.001-0.2%; and excipients 40-50%.

In some embodiments, the red yeast rice is one or more of red yeast rice powder, red yeast rice extract, and functional red yeast rice. In some embodiments, the red yeast rice is functional red yeast rice. In some embodiments, the weight percentage of lovastatin in the red yeast rice is about 1-5%, preferably about 3%. In some embodiments, the red yeast rice is functional red yeast rice, wherein the weight percentage of lovastatin is about 1-5%, preferably about 3%.

In some embodiments, the weight percent content of total flavonol glycosides in the Ginkgo biloba extract is 24-30%, preferably about 24%. In some embodiments, the extraction method of the Ginkgo biloba extract comprises crushing the Ginkgo biloba leaves, adding 3-5 times dilute ethanol (for example, 50vol.% ethanol) and refluxing extraction at 70-80°C for 3 times, 2-3 times each time. Hours, adsorbed with macroporous resin, eluted with water and different concentrations of ethanol.

In some embodiments, the weight ratio of the functional red yeast rice to the Ginkgo biloba extract is about 2-3:1, such as 2:1, 2.1:1, 2.2:1, 2.3:1, 2.4:1 , 2.5:1, 2.6:1, 2.7:1, 2.8:1, 2.9:1 or 3:1.

In some embodiments, the health care composition for regulating blood lipids comprises, by weight percentage: 38.9% of red yeast rice; 16.7% of ginkgo biloba extract; 0.00067% of folic acid; 0.0011% of vitamin B6; and 44.3% of auxiliary materials.

In some embodiments, the excipients include fillers and lubricants. In some embodiments, the filler comprises one or more of maltodextrin, microcrystalline cellulose, and lactose. In some embodiments, the lubricant comprises silica. In some embodiments, the excipients comprise maltodextrin, microcrystalline cellulose, lactose and silicon dioxide.

In some embodiments, the health care composition comprises 140 mg of functional red yeast rice powder based on 0.36 g. In some embodiments, the health care composition comprises 60 mg of Ginkgo biloba extract based on 0.36 g. In some embodiments, the health care composition comprises 0.24 mg folic acid based on 0.36 g. In some embodiments, the health care composition comprises 0.4 mg vitamin B6 based on 0.36 g. In some embodiments, the health care composition comprises 140 mg of functional red yeast rice powder, 60 mg of ginkgo biloba extract, 0.24 mg of folic acid and 0.4 mg of vitamin B6 based on 0.36 g. In some embodiments, the health care composition comprises 140 mg of functional red yeast rice powder, 60 mg of ginkgo biloba extract, 0.24 mg of folic acid, 0.4 mg of vitamin B6, 73.76 mg of maltodextrin, 46 mg of microcrystalline cellulose, Lactose 36mg and Silicon Dioxide 3.6mg.

In some embodiments, the health care composition comprises 4.2 mg of lovastatin based on 0.36 g. In some embodiments, the health care composition comprises 7.2 mg of total flavonol glycosides of Ginkgo biloba based on 0.36 g. In some embodiments, the health care composition comprises 4.2 mg of lovastatin, 7.2 mg of total ginkgo flavonol glycosides, 0.24 mg of folic acid and 0.4 mg of vitamin B6 based on 0.36 g.

In some embodiments, the Ginkgo biloba extract can be replaced by Ginkgo biloba. If replace Ginkgo biloba extract with Ginkgo biloba, then the weight of Ginkgo biloba should be 50-100 times of the weight of Ginkgo biloba extract used, preferably 60 times, and the content of red yeast rice, folic acid, vitamin B6 in the composition can remain unchanged. For example, if 3600 mg of Ginkgo biloba extract is used instead of 60 mg of Ginkgo biloba extract, the single dose of the prepared health care composition will increase accordingly. Correspondingly, for example, the health care composition contains, by weight percentage: 3.6% of red yeast rice; 92.3% of ginkgo biloba; 0.0007% of folic acid; 0.001% of vitamin B6; and 4.01% of auxiliary materials. Correspondingly, the weight ratio of red yeast rice and ginkgo biloba in the health care composition is, for example, about 1:20-30, such as 1:21, 1:22, 1:23, 1:24, 1:25, 1:26, 1 :27, 1:28, 1:29, or 1:30.

In the present application, by adding folic acid and vitamin B6 to the composition of red yeast rice and ginkgo leaf extract, the inventors unexpectedly found that it not only can significantly reduce The levels of serum TG, TC, LDL-C, and can significantly increase the level of HDL-C, so that blood lipids can be more effectively regulated, and used to prevent various cardiovascular diseases caused by dyslipidemia, including but not limited to simple serum Hypercholesterolemia with elevated TC, hypertriglyceridemia with elevated serum TG alone, high LDL-C hyperlipidemia with high serum LDL-C level, low HDL-C with low serum HDL-C level alone C blood, two or all of the three serum TC, TG, LDL-C elevated mixed hyperlipidemia, two or all of the serum TC, TG, LDL-C elevated combined serum Mixed hyperlipidemia, xanthoma, atherosclerosis, coronary heart disease and peripheral vascular disease with low HDL-C.

Description of drawings

Fig. 1 is the change of serum high-density lipoprotein cholesterol (HDL-C) level of rats after different treatment groups. (A) concentration of serum HDL-C of each group of rats before taking medicine; (B) concentration of serum HDL-C of each group of rats after taking medicine; (C) mean value of concentration of serum HDL-C of each group of rats. *, P<0.05; ***, P<0.001; n.s., no significant difference; n=12/group.

Fig. 2 is the change of rat serum total cholesterol (TC) level after different treatment groups. (A) The concentration of serum TC in each group of rats before taking medicine; (B) The concentration of serum TC in each group of rats after taking medicine; (C) The concentration mean value of serum TC in each group of rats. ***, P<0.001; n.s., no significant difference; n=12/group.

Fig. 3 is the change of serum low-density lipoprotein cholesterol (LDL-C) level of rats after different treatment groups. (A) The concentration of serum LDL-C in each group of rats before taking the medicine; (B) The concentration of serum LDL-C in each group of rats after taking the medicine; (C) The mean value of the concentration of serum LDL-C in each group of rats. ***, P<0.001; n.s., no significant difference; n=12/group.

Fig. 4 is the change of serum triglyceride (TG) level of rats after different treatment groups. (A) The concentration of serum TG in each group of rats before taking medicine; (B) The concentration of serum TG in each group of rats after taking medicine; (C) The concentration mean value of serum TG in each group of rats. ***, P<0.001; n.s., no significant difference; n=12/group.

Fig. 5 is a representative picture of HE staining pathological examination of various organs of acute toxicity experiment mice (A) and rats (B).

detailed description

The inventors of the present application found that adding an appropriate amount of folic acid and vitamin B6 to lipid-lowering health products containing red yeast rice and ginkgo biloba can significantly increase the level of HDL-C while significantly reducing the levels of serum TG, TC, and LDL-C . This effect is unexpected, because on the one hand it is known in the prior art that Ginkgo biloba or its extracts can only be used at very high doses (corresponding to rat doses of 1.0 g/kg and 2.0 g/kg, converted to adult doses of about 12g/60kg body weight or 24g/60kg body weight) can have the effect of raising HDL-C, but at low doses (corresponding to the rat dose of 0.5g/kg, converted to an adult dose of about 6g/60kg body weight) there is no such effect, However, the daily dose of ginkgo extract in health care products in my country must not exceed 200 mg, which causes the effect of ginkgo extract to increase HDL-C and cannot be reflected at the prescribed dose of health care products; The additional folic acid and vitamin B6 are known not to have the effect of raising HDL-C, and before the present invention was made, it was expected that the combination of the two with red yeast rice and ginkgo biloba would not raise HDL-C. As mentioned earlier, raising HDL-C has significant health benefits in subjects with dyslipidemia, especially low HDL-C.

As demonstrated by the following experimental examples, the health care composition of the present invention can significantly increase HDL-C levels at low, medium and high doses, while significantly reducing serum TG, TC, and LDL-C levels. The three doses are 0.03g/kg, 0.06g/kg, and 0.18g/kg, respectively, and the adult doses are about 0.36g/60kg, 0.72g/60kg, and 2.16g/60kg. Compared with the prior art, the low dose of the composition of the present invention is as low as about 6% of the above-mentioned Yinqu capsule low-dose group and about 3% of the above-mentioned Yinqu capsule medium-dose group, but it can still significantly reduce serum TG and TC , LDL-C levels, and significantly increase HDL-C levels. When the health-care composition of the present invention was used in a much lower dose, the usage amount of its component GBE would also be significantly reduced, thereby eliminating the possible side effects caused by high-dose GBE, but its effect of increasing HDL-C levels was not not eliminated. In addition, adding folic acid and vitamin B6 to the health care composition can bring additional health benefits.

health care composition

The health care composition provided by the application can be used to regulate blood lipids, including significantly reducing serum TG, significantly reducing TC, significantly reducing LDL-C and/or significantly increasing HDL-C. The health-care composition can be used for subjects with dyslipidemia, such as hypercholesterolemia with elevated serum TC, hypertriglyceridemia with elevated serum TG, and high LDL-C blood with excessively high serum LDL-C levels. Hyperlipidemia, hypo-HDL-C syndrome with low serum HDL-C level, mixed hyperlipidemia with elevated serum TC, TG, LDL-C, two or all of them, serum TC, TG, LDL -Mixed hyperlipidemia with elevated levels of two or all of the three combined with low serum HDL-C. The health care composition can also be used to prevent various cardiovascular diseases caused by dyslipidemia, including but not limited to xanthoma, atherosclerosis, coronary heart disease and peripheral vascular disease. The increase (too high) or decrease (too low) is relative to the normal value, and the range of increase or decrease is not limited. The reference normal values of each blood lipid are as follows TC: <5.20mmol/l; TG: <1.70mmol/l; HDL-C: >0.91mmol/l; LDL-C: <3.12mmol/l.

In the present invention, the "significantly increased HDL-C", "significantly increased HDL-C" or similar terms mean that the composition of the present invention has a statistically significant Differentially elevated levels of HDL-C. Similarly, said "significantly lowering" or similar terms means that the composition of the present invention lowers TC, TG or LDL-C by a statistically significant difference compared to a subject whose TC, TG or LDL-C is lowered or is too low s level. In some embodiments, "statistically significant difference" means that if the data satisfies the normal distribution, one-way analysis of variance (One-Way ANOVA) is used to analyze the data and compare the differences between groups; if the data does not satisfy the normality or homogeneity of variance, non-parametric test (Kruskal-Wallis rank sum test) was used for data analysis and comparison of differences between groups; P<0.05.

In some embodiments, the present application provides a health care composition for regulating blood lipids, comprising: 30-46% of red yeast rice; 10-28% of ginkgo biloba extract; 0.0005-0.056% of folic acid; Vitamin B6 0.001-0.2%; and accessories 40-50%.

In the present application, the red yeast rice can be one or more of red yeast rice powder, red yeast rice extract, and functional red yeast rice, preferably functional red yeast rice. In a preferred embodiment, the weight percentage of lovastatin in red yeast rice is about 1-5%, preferably about 3%, and the lovastatin can be a mixture of its acid form and lactone form . Red yeast rice can be prepared by conventional methods or commercially available. Monascus is produced by the mycelium of the Aspergillus family fungus Monascus purpureus Went. parasitizing on rice (such as japonica rice). Monascus is aliased in Chinese as red yeast rice, red rice, and red rice. It is a genus of Ascomycetes fungi in the family of Aspergillus. , Rusty Monascus Berry, Red Monascus. This application preferably uses the red yeast rice fermented with Monascus purpura, and the representative source is the functional red yeast rice powder produced by Hangzhou Shuangma Biotechnology Co., Ltd.

In some embodiments, the weight percent content of total flavonol glycosides in the Ginkgo biloba extract is 24-30%, preferably about 24%. In some embodiments, the extraction method of the Ginkgo biloba extract comprises crushing the Ginkgo biloba leaves, adding 3-5 times dilute ethanol (for example, 50vol.% ethanol) and refluxing extraction at 70-80°C for 3 times, 2-3 times each time. Hours, adsorbed with macroporous resin, eluted with water and different concentrations of ethanol. The present application expects that the Ginkgo biloba leaf extract with a total flavonol glycoside content ≥ 24% extracted by other methods or commercially available can be used in accordance with the Pharmacopoeia standard.

In one embodiment, the health care composition comprises 140 mg of functional red yeast rice powder, 60 mg of ginkgo biloba extract, 0.24 mg of folic acid and 0.4 mg of vitamin B6 based on 0.36 g. In one embodiment, the health care composition comprises 140mg functional red yeast rice powder, 60mg ginkgo biloba extract, 0.24mg folic acid, 0.4mg vitamin B6, 73.76mg maltodextrin, 46mg microcrystalline cellulose, Lactose 36mg and Silicon Dioxide 3.6mg.

In one embodiment, the health care composition comprises 4.2 mg of lovastatin, 7.2 mg of total flavonol glycosides of Ginkgo biloba, 0.24 mg of folic acid and 0.4 mg of vitamin B6 based on 0.36 g.

In one embodiment, the Ginkgo biloba extract can be replaced by Ginkgo biloba. Correspondingly, for example, the health care composition contains, by weight percentage: 3.6% of red yeast rice; 92.3% of ginkgo biloba; 0.0007% of folic acid; 0.001% of vitamin B6; and 4.01% of auxiliary materials.

The content limit of each ingredient in health care products in my country is stipulated as follows: lovastatin 10mg/d, GBE 200mg/d (converted to total flavonol glycosides is 48mg/d), folic acid 0.4mg/d, vitamin B6 1.5mg/d. In one embodiment, the health care composition is made into a unit dose of 0.36 g, and the dosage form can be capsules, tablets, granules, powders or other suitable dosage forms. The preparation method of each dosage form refers to the conventional method operation in the field of pharmacy. In one embodiment, the daily dosage of the health care composition is 0.72g.

Example

Embodiment 1. The preparation method of health care composition capsule

According to the formula of 1000 capsules: 140g functional red yeast rice powder, 60g ginkgo biloba extract, 0.24g folic acid, 0.4g vitamin B6, 73.76g maltodextrin, 46g microcrystalline cellulose, 36g lactose and 3.6g silicon dioxide, first Mix maltodextrin, folic acid and vitamin B6 uniformly in equal increments, then put red yeast rice, ginkgo biloba extract, lactose and the uniformly mixed maltodextrin, folic acid and vitamin B6 into a wet mixing granulator , after mixing for 30 minutes, add an appropriate amount of purified water, pass through a 60-mesh sieve and granulate. The obtained wet granules are dried until the water content is less than or equal to 5.0%, passed through a 60-mesh sieve for sizing, added microcrystalline cellulose and silicon dioxide, mixed in a mixer for 30 minutes, filled into capsules, and obtained capsules.

Embodiment 2. the preparation method of health care composition tablet

With the formula described in Example 1, mix maltodextrin, folic acid and vitamin B6 uniformly in an equal increasing method, and put red yeast rice, ginkgo biloba extract, lactose and maltodextrin, folic acid and vitamin B6 In a wet mixing granulator, add an appropriate amount of purified water after mixing for 30 minutes, pass through a 60-mesh sieve for granulation, dry the obtained wet granules until the moisture content is ≤ 5.0%, pass through a 60-mesh sieve for granulation, add microcrystalline cellulose, di The silicon oxide was mixed in a mixer for 30 minutes, and compressed into tablets to obtain tablets.

Embodiment 3. the preparation method of health care composition granule

Using the formula described in Example 1, first mix the maltodextrin, folic acid and vitamin B6 uniformly in an equal increasing method, and then mix the red yeast rice, ginkgo leaf extract, lactose and the uniformly mixed maltodextrin, folic acid and vitamin B6 Place in a wet mixing granulator, mix for 30 minutes, add an appropriate amount of purified water, pass through a 60-mesh sieve and granulate. The obtained wet granules are dried until the water content is less than or equal to 5.0%, passed through a 60-mesh sieve for sizing, adding microcrystalline cellulose and silicon dioxide, mixing in a mixer for 30 minutes, and bagging to obtain granules.

Embodiment 4. the preparation method of health care composition powder

With the formula described in Example 1, pass maltodextrin, folic acid and vitamin B6 through a 100-mesh sieve respectively, weigh maltodextrin, folic acid and vitamin B6 according to the formula amount, and mix them uniformly in an equal increment method. Leaf extract, lactose, microcrystalline cellulose, and silicon dioxide are respectively passed through a 100-mesh sieve, and the sieved red yeast rice, ginkgo biloba extract, lactose, microcrystalline cellulose, silicon dioxide are weighed according to the formula and mixed evenly The maltodextrin, folic acid and vitamin B6 were put into a mixer and mixed for 30 minutes to obtain a powder.

Experimental example

experimental method

Preparation of animal model of hyperlipidemia and grouping of animals: 12 rats were given maintenance diet as blank control group, and 60 rats were given high-fat diet to prepare animal model of hyperlipidemia. After feeding the high-fat diet for 14 days, the rats in the blank control group and the rats fed with the high-fat diet were not fasted to take blood, and the blood was collected and centrifuged to separate the serum, and then the serum total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol and Triglyceride level, according to the results to determine whether the model is successfully made. According to the total cholesterol level, the rats fed with high-fat diet were further randomly divided into five groups with 12 rats in each group, namely model control group, low-dose ST group (low-dose ST group), middle-dose group (medium-dose ST group), high-dose ST group, Dose group (high-dose ST group) and drug control group (red yeast powder and ginkgo leaf extract powder mixed by weight 1:1). Animals in each dose ST group and drug control group were fed with high-fat feed, and were given the corresponding ST or drugs by intragastric administration once a day; Bacterial water was given once; while the animals in the blank control group were given maintenance feed, they were given sterile water by gavage once a day. After gavage for 45 days, the blood was collected without fasting and the serum was separated to measure total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol and triglyceride in the serum of rats in each group.

Dosage and administration method: According to the recommended dose of the capsule is 0.72g/60kg per day for the human body, the low, medium, and high doses of the ST group used in rat animal experiments are 2.5 times, 5 times, and 15 times the recommended amount for the human body. Times, that is, 0.03g/kg, 0.06g/kg, 0.18g/kg per day. The preparation method and administration method of the ST samples used in the experiment of each dosage group are as follows. The ST low-dose group took 0.30 g of the capsule content of Example 1, added sterile water to 50.0 mL; the ST middle-dose group took 0.60 g of the capsule content of Example 1, added sterile water to 50.0 mL; the ST high-dose group took the Example 1 Capsule content 1.80g, add sterile water to 50.0mL. The dose of the drug control group was 0.060g/kg, 0.3g of red yeast rice powder (60 mesh) and 0.3g of ginkgo leaf extract powder (60 mesh) were taken, and sterile water was added to 50.0mL. Rats were given intragastric administration at 5 mL/kg once a day for 45 consecutive days, and serum was collected and separated for determination of various indicators. Blank control group (0g/kg) and model control group (0g/kg) were administrated with sterile water at 5mL/kg daily.

Determination of serum total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol and triglycerides: take venous blood, centrifuge at 3000r/min for 10 minutes to separate serum, and use an automatic biochemical analyzer to measure serum total cholesterol and high-density lipoprotein Protein cholesterol, LDL cholesterol and triglycerides.

Data processing: Graphpad Prism 9.0 software was used for data statistical analysis and chart making, and the data were expressed as mean ± standard deviation (Mean ± SD). If the data satisfies normal distribution and homogeneity of variance, one-way analysis of variance (One-Way ANOVA) was used for data analysis and comparison of differences between groups; if the data did not meet normal distribution or homogeneity of variance, nonparametric test (Kruskal -Wallis rank sum test) for data analysis and comparison of differences between groups. P<0.05 means that the difference between groups is statistically significant.

Experimental Example 1. The lipid-lowering composition of the present invention significantly increases serum high-density lipoprotein cholesterol (HDL-C) level, while the control drug had no such effect

As shown in Figure 1A, after the blank control group was fed with maintenance feed and other experimental animals were fed with high-fat feed for 14 days, compared with the blank control group, the serum HDL-C levels of rats in each group fed with high-fat feed (average value ranged from 0.73 to 0.76mmol/L) were significantly lower than the blank control group (mean 0.92mmol/L), indicating that high-fat feeding caused a significant decline in rat serum HDL-C levels.

As shown in Figure 1B, after feeding the blank control group with maintenance diet and other experimental animals fed with high-fat diet and given different drugs for 45 days, the serum HDL-C level (0.47±0.08mmol/L) of the rats in the model control group was significantly increased. Lower than the blank control group (0.90±0.08mmol/L), while the low, medium and high dose ST group rat serum HDL-C levels (respectively 0.55±0.08, 0.54±0.11 and 0.55±0.10mmol/L) were significantly Higher than the model control group (0.47 ± 0.08mmol/L), showing that each dose of ST all significantly improves the rat serum HDL-C level; and the serum HDL-C level of the drug control group rats (0.50 ± 0.07mmol/L) and There was no significant difference compared with the model control group. The above results show that Tiaozhi Capsules can significantly increase HDL-C, while the control drug has no such effect.

Experimental Example 2. The lipid-lowering composition of the present invention significantly reduces serum total cholesterol (TC)

As shown in Figure 2A, after the blank control group was fed with maintenance diet and other experimental animals were fed with high-fat diet for 14 days, compared with the blank control group, the TC levels of the rats in each group fed with high-fat diet (mean between 3.31 and 3.43mmol/ L) were significantly higher than that of the blank control group (the average value was 2.35mmol/L), indicating that the hyperlipidemia animal model was successfully established.

As shown in Figure 2B, after the blank control group was fed with maintenance feed and other experimental animals were fed with high-fat feed and given different drugs for 45 days, the serum TC level (4.24±0.45mmol/L) of the rats in the model control group was significantly higher In the blank control group (1.76 ± 0.27mmol/L), compared with the model control group, the serum TC levels of the rats in the low, medium and high dose ST groups and the drug control group (respectively 2.33 ± 0.40, 2.33 ± 0.48, 2.38 ±0.49 and 2.19±0.33mmol/L) were significantly lower than the model control group (4.24±0.45mmol/L). The above results showed that each dose of ST and the control drug significantly reduced the level of rat serum total cholesterol TC.

Experimental Example 3. The lipid-lowering composition of the present invention significantly reduces serum low-density lipoprotein cholesterol (LDL-C)

As shown in Figure 3A, after the blank control group was fed with maintenance diet and other experimental animals were fed with high-fat diet for 14 days, compared with the blank control group, the serum LDL-C levels of rats in each group fed with high-fat diet (average value ranged from 1.67 to 1.71 mmol/L) were significantly higher than the blank control group (the mean was 0.20mmol/L), indicating that the hyperlipidemia animal model was successfully established.

As shown in Figure 3B, after feeding the blank control group with maintenance diet and other experimental animals fed with high-fat diet and given different drugs for 45 days, the serum LDL-C level (1.62±0.25mmol/L) of the rats in the model control group was significantly increased. Higher than the blank control group (0.17 ± 0.03mmol / L), and compared with the model control group, low, medium and high dose ST group and drug control group rat serum LDL-C levels (respectively 0.54 ± 0.14, 0.53 ± 0.12, 0.54±0.15 and 0.55±0.13mmol/L) were significantly lower than the model control group (1.62±0.25mmol/L). The above results showed that each dose of ST and control drugs could significantly reduce the level of serum low-density lipoprotein cholesterol LDL-C in rats.

Experimental Example 4. The lipid-lowering composition of the present invention significantly reduces serum triglycerides (TG)

As shown in Figure 4A, after the blank control group was fed with maintenance diet and other experimental animals were fed with high-fat diet for 14 days, compared with the blank control group, the serum TG levels of rats in each group fed with high-fat diet (mean value was between 5.37 and 5.84mmol/ L) were significantly higher than the blank control group (mean value was 2.70mmol/L), indicating that the hyperlipidemia animal model was successfully established.

As shown in Figure 4B, after feeding the blank control group with maintenance diet and other experimental animals fed with high-fat diet and given different drugs for 45 days, the serum TG level (3.20±0.47 mmol/L) of the rats in the model control group was significantly higher than that of Blank control group (1.50 ± 0.36mmol/L), compared with the model control group, low, medium and high dose ST group and drug control group rat serum TG levels (respectively 2.26 ± 0.55, 2.26 ± 0.65, 2.24 ± 0.78 and 1.61±0.39mmol/L) were significantly lower than the model control group (3.20±0.47mmol/L). The above results showed that each dose of ST and the control drug significantly reduced the level of serum triglyceride TG in rats.

The above results show that the lipid-lowering composition of the present invention can reduce serum total cholesterol, low-density lipoprotein cholesterol and triglyceride levels, and has an obvious blood-lipid-lowering effect. The lipid-regulating composition of the present invention can also improve the level of high-density lipoprotein cholesterol, while the drug control has no effect on the high-density lipoprotein cholesterol, indicating that the lipid-regulating composition of the present invention is better than the control drug in terms of improving high-density lipoprotein cholesterol.

Experimental example 5. Acute toxicity test did not find ST-induced heart, liver, and kidney damage lesions

Grouping and administration: referring to the acute toxicity test method in the Implementation Manual of Health Food Inspection and Technical Regulations, the safety of the powder composition ("ST") of Example 4 was evaluated by the maximum tolerated dose method. Forty BALB/c mice and 10 SD rats were randomly divided into four groups, half male and half male. There were 20 mice in the normal control group and 20 in the ST group; 4 rats in the normal control group and 6 in the ST group. Animals were fasted overnight before administration, the concentration of ST solution was 0.58g/mL, and the volume of intragastric administration was 20mL/kg body weight. The administration was administered twice a day with an interval of 4h, and the total administration dose was 23.2g/kg/d. Since the dosage of ST human is 0.012g/kg/d, the dosage of this rat is 1933 times of human dosage. The animals were weighed before and after the administration, and the death and poisoning manifestations of the animals were observed and recorded within 72 hours; at 72 hours, the hearts, livers and kidneys of the animals in each group were taken for pathological examination.

HE staining method: Take the tissue out of the fixative, soak in 50% ethanol (30min) - 70% ethanol (overnight) - 80% ethanol (30min) - 90% ethanol (30min) - 95% ethanol (30min) - none Water ethanol (2 times, 30 min each time) - xylene (2 times, 5-10 min each time, until the sample is completely transparent) - 62°C paraffin (3 times, 1 h each time), and then tissue embedding. Tissue paraffin sections with a thickness of 3 μm were dried on a drying machine and dried in a 37°C oven overnight. Then, the paraffin sections stored at room temperature were taken out and baked in a 65°C oven for 30 minutes, and then immediately immersed in two Dewax in toluene for three times, each time for 5 minutes; the rehydration process is the third soaking in xylene, then soak the slices in 100% ethanol-100% ethanol-95% ethanol-90% ethanol-80% ethanol-70% Ethanol-50% ethanol-distilled water for rehydration, 1 min each time; take out the slices to dry slightly, then place the slices in a wet box, add hematoxylin staining solution on the tissue to ensure that the staining solution completely covers the tissue, and incubate at room temperature for 5 min ; Gently rinse the slices with distilled water to remove excess hematoxylin, then put the slices back into the wet box, add eosin staining solution on the tissue, and incubate at room temperature for 2 minutes; gently rinse the slices with distilled water; soak the slices in sequence 90% ethanol (1min)-95% ethanol (1min)-100% ethanol (1min)-100% ethanol (1min)-xylene (5min)-xylene (5min) for dehydration and transparency, and then use neutral resin (with Appropriate amount of xylene diluted, about 50% xylene) for mounting. ECLIPSE Ci-S upright microscope was used to take bright-field photos, and each individual randomly selected a field of view to take photos for pathological analysis.

Data statistics: Data are expressed as mean ± standard deviation (Mean ± SD). Graphpad Prism 7.0 software was used to analyze and graph the data. If the data satisfied normality and homogeneity of variance, one-way analysis of variance (One-Way ANOVA) was used, and Fisher's LSD test was used for comparison between groups. If the data did not satisfy normality or homogeneity of variance, a non-parametric test (Kruskal-Wallis rank sum test) was used.

General clinical observation and body weight

In the acute toxicity test, no animal died in the administration group within 72 hours after 2 gavages, and the behaviors and activities of small and medium-sized rats were normal, and there was no obvious poisoning reaction. After 72 hours of anatomy, the heart, liver, kidney and other major organs were observed with the naked eye, and no abnormal color and shape were found, nor were there any bleeding points or other pathological changes. Compared with the control group, there was no significant difference (P>0.05) between the animal body weight and the coefficient of each organ in each group before and after administration (data not shown).

Acute toxicity test results show that the maximum tolerated dose (Maximal Tolerable Dose, MTD) of the composition of the present invention is 23.2 g/kg, equivalent to 1933 times of human oral dose, and LD ₅₀ cannot be measured. Classified according to acute toxicity dose, it belongs to non-toxic grade.

Histopathological observation

The main organs such as the heart, liver, and kidney of mice and rats were not abnormal in color and shape, and there were no bleeding points or other pathological changes, so there was no abnormality. Compared with the normal group, the HE staining results showed that acute administration of the composition of the present invention did not see pathological damage to the main organs of the heart, liver, and kidney of mice and rats, and the structures of each organ were clear and the cell morphology was normal (see Figure 5) The data show that the composition of the present invention is safe on the basis of 1933 times higher than the human recommended dose.

In this experiment, the maximum tolerated dose method was used to evaluate the acute toxicity of the composition of the present invention to two rodents, BALB/c mice and SD rats. Acute toxicity test result shows that composition of the present invention is 23.2g/kg/d to the maximum tolerated dose (Maximal Tolerable Dose, MTD) of mouse, is equivalent to 1933 times of people's oral dose, observes under this dose The results of pathological sections of various organs of mice and rats showed no damage to the heart, liver, and kidney. Classified according to the acute toxicity dose, it belongs to the non-toxic grade, indicating that it is safe to eat according to the recommended amount.

Claims

A health care composition for raising HDL-C, comprising: 30-46% of red yeast rice; 10-28% of ginkgo biloba extract; 0.0005-0.056% of folic acid; 0.001-0.2% of vitamin B6; and 40-50% of auxiliary materials, wherein the health care composition significantly reduces the levels of serum TG, TC, and LDL-C, and at the same time significantly increases the level of HDL-C.
The health care composition according to claim 1, wherein the red yeast rice is one or more of red yeast rice powder, red yeast rice extract, and functional red yeast rice.
The health care composition according to claim 1, wherein the red yeast rice is functional red yeast rice, and the weight percentage of lovastatin in the functional red yeast rice is 1-5%.
The health care composition according to claim 3, wherein the weight percentage of lovastatin in the functional red yeast rice is 3%.
The health care composition according to claim 1, wherein the weight percentage of total flavonol glycosides of the Ginkgo biloba extract is 24-30%.
The health care composition according to claim 5, wherein the weight percentage of the total flavonol glycosides of the Ginkgo biloba extract is 24%.
The health-care composition according to claim 1, wherein the health-care composition for regulating blood lipids comprises, by weight percentage: 38.9% of red yeast rice; 16.7% of ginkgo biloba extract; 0.00067% of folic acid; 0.0011% of vitamin B6 and excipients 44.3%.
The health care composition according to claim 1, wherein the health care composition comprises 140mg functional red yeast rice powder, 60mg ginkgo leaf extract, 0.24mg folic acid and 0.4mg vitamin B6 in 0.36g.
The healthcare composition according to claim 1, wherein the adjuvants comprise fillers and lubricants.
The health care composition according to claim 9, wherein the filler comprises one or more of maltodextrin, microcrystalline cellulose and lactose, and the lubricant comprises silicon dioxide.
The health-care composition according to claim 10, wherein the health-care composition comprises 140mg functional red yeast rice powder, 60mg ginkgo biloba extract, 0.24mg folic acid, 0.4mg vitamin B6, 73.76mg maltodextrin by 0.36g, Microcrystalline Cellulose 46mg, Lactose 36mg and Silicon Dioxide 3.6mg.
The health care composition according to claim 1, wherein the health care composition comprises 4.2 mg lovastatin, 7.2 mg total flavonol glycosides of Ginkgo biloba, 0.24 mg folic acid and 0.4 mg vitamin B6 in 0.36 g.
A health care composition for raising HDL-C, comprising: 3.6% of red yeast rice; 92.3% of ginkgo biloba; 0.0007% of folic acid; 0.001% of vitamin B6; and 4.01% of auxiliary materials; The composition significantly reduces the levels of serum TG, TC and LDL-C, and at the same time significantly increases the level of HDL-C.