CN103585234B - The application of Radix Berberidis Amurensis berry extract in preparation treatment diabetes medicament - Google Patents

Abstract

The invention discloses a new application of the berberis fruit extract, which is the application of the berberis fruit extract in the preparation of medicines for treating diabetes. The extract is prepared according to the following method: using barberry fruit as raw material, extracting with water, and purifying through two macroporous adsorption resins, including berberry fruit extract A and barberry fruit extract B. The barberry fruit extract provided by the invention is of great significance for the preparation of medicines for treating diabetes.

Description

Application of berberis fruit extract in the preparation of medicines for treating diabetes

technical field

The invention belongs to the field of biomedicine, and relates to the application of an extract of Chinese medicinal materials in the preparation of medicines for treating diabetes, in particular to the application of extracts of berberis fruit in the preparation of medicines for treating diabetes.

Background technique

With the aging of the world's population, diabetes has become a common and frequently-occurring disease, and its incidence is on the rise in industrialized countries. According to statistics, there are about 150 million diabetic patients in the world, 80% of which are non-insulin-dependent diabetes (type 2), of which there are about 30 million in my country. The number of deaths caused by diabetes complications ranks third after cardiovascular and cerebrovascular diseases and cancer in developed countries, which has attracted great attention from all over the world, and researches on the pathogenesis and drug prevention and treatment of diabetes have been carried out one after another. Since the etiology of type 1 diabetes is basically clear, only insulin supplementation is enough, while type 2 diabetes is more complicated. At present, the medical field in my country adopts comprehensive treatment measures including diet therapy, drug therapy and other adjuvant therapies. Although western medicine emerges in an endless stream, it cannot fundamentally solve the problem, and western medicine treatment has certain side effects and "treatment failure". TCM treatment of diabetes has a long history, rich clinical experience, and remarkable curative effect. It has become a research hotspot at home and abroad in recent years. The understanding of diabetes in traditional Chinese medicine is different from that in western medicine. Traditional Chinese medicine emphasizes the holistic view and reflects the essence of the disease through the changes of yin and yang. The normal body is "yin and yang secret", yin and yang are balanced, that is, the internal environment of the body maintains a stable state, and the glucose metabolism also maintains stability; and the toxic and side effects of traditional Chinese medicine are relatively small, and patients can take it for a long time. This is a unique advantage that western medicine cannot match. .

Traditional Chinese medicine believes that diabetes is caused by burning yin deficiency in the lungs, stomach, and kidneys, and abnormal water and grain transfer. Its base

The pathogenesis is depletion of yin and fluid, excess of dryness and heat, deficiency of kidney yin as the root, dryness and heat of the lung and stomach as the standard, long-term illness leads to loss of yin, depletion of qi and injury of yang, resulting in damage to both qi and yin, deficiency of both yin and yang, blood stasis in the veins, The tendons and arteries are not nourished, the internal organs are damaged, and a series of concurrent diseases gradually appear.

The course of diabetes is a process of yin damage and yang. Based on yin differentiation, from mild to severe, it is the transformation of yin deficiency - deficiency of both qi and yin - deficiency of both yin and yang. And blood stasis runs through the whole process of primary disease and concurrent disease.

In recent years, some scholars have explored the relationship between TCM syndrome differentiation and clinical biochemical indicators of diabetes. From the changes of pancreatic islet function, blood lipid level, plasma sex hormone, corticosteroid level, immune function and blood rheology, etc., observe the correlation between the type of diabetes and the changes of the above indicators, and find the abnormal changes of various indicators, ranging from mild to severe , mostly develops in the direction of deficiency of yin—deficiency of both qi and yin—deficiency of both yin and yang. It embodies the material basis for exploring the classification of diabetes by combining traditional Chinese and Western medicine.

Syndrome differentiation and treatment is the essence of TCM theory. According to the main pathogenesis of diabetes, traditional Chinese medicine distinguishes the severity of dryness and heat and yin deficiency, and pays attention to nourishing yin when taking medicine. If there is dryness and heat, it should be cleared away. According to modern pharmacological concepts, the research on the anti-diabetic effect of traditional Chinese medicine is mainly as follows. Single traditional Chinese medicine: coptis and berberine, bitter gourd, ginseng, astragalus, scutellaria baicalensis, mulberry leaves, traditional Chinese medicine polysaccharides (such as tremella polysaccharides, tremella spore polysaccharides, fungus polysaccharides, Hericium erinaceus polysaccharides, laver polysaccharides, Ophiopogon japonicus polysaccharides, laminarin ), Lycium barbarum polysaccharide-X, Lycium barbarum polysaccharide-D, papaya polysaccharide and paeonol polysaccharide, etc.), silymarin, rhubarb. In addition, oral administration of flavonoids puerarin or clausenacoumarine also reduced hyperglycemia in ALX animals. The basic principles of TCM treatment of diabetes can be summarized as: nourishing yin and clearing away heat, nourishing qi and nourishing yin, nourishing kidney and spleen, promoting blood circulation and removing blood stasis, etc. Commonly used classic prescriptions include Shenbaihu Decoction, Liuwei Dihuang Pills, Bawei Dihuang Pills, and Qianjin Huanglian Pills. According to different syndrome types and the above-mentioned treatment principles, formulas can be formulated or added or subtracted from ancient prescriptions.

Berberis berry is the dry and mature fruit of Berberis nummularia Bge. Included in the 1998 "Ministry of Health of the People's Republic of China Drug Standard Uighur Medicine Sub-volume". It is flat in nature, has the effects of invigorating the stomach and harmonizing the middle, promoting body fluid and quenching thirst, clearing away heat and detoxifying. For dyspepsia, diarrhea, thirst, aphtha, pharyngitis. It is a commonly used drug in ethnic minority areas such as Tibet and Mongolia. However, the basic research on Berberis fruit is still very limited, which limits the subsequent promotion and application of this medicinal material. Modern research shows that berberberry fruit contains alkaloid components such as berberine, jatrorrhizine, and palmatine, as well as anthocyanins such as geranium pigment and mallow pigment. Other chemical components are unknown; no pharmacological studies have been reported.

According to domestic patent search results, there is no patent related to barberry fruit.

In the above-mentioned documents and patents, there is no report on the use of Berberis fruit extract for the preparation of drugs for treating diabetes.

Contents of the invention

The object of the present invention is to provide the application of barberry fruit extract in the preparation of medicines for treating diabetes.

The present invention is achieved through the following technical solutions:

The barberry fruit used in the present invention is the dry mature fruit of Berberis nummularia Bge.

The application of the Berberis fruit extract of the present invention in the preparation of medicines for treating diabetes, the preparation method of the Berberis fruit extract is:

(1) Berberis fruit, water is used as solvent, the extraction temperature is 50°C-95°C, the number of extractions is 1-3 times, each extraction time is 1-4 hours, and the amount of each solvent is 6-15% of the weight of Berberis fruit times; filter, combine extracts, concentrate to relative density d=1.10-1.18, filter to obtain liquid A;

(2) Pass the medicinal liquid A obtained in step (1) through a non-polar macroporous adsorption resin, and first elute with water, and the water eluate directly passes through the polar macroporous adsorption resin, and then use a Elute the non-polar macroporous adsorption resin with ethanol solution, collect ethanol eluents with different concentrations, concentrate and dry to obtain Berberis fruit extract A; then elute the polar macroporous adsorption resin with ethanol solution with a concentration of 50%-95% Resin, collect ethanol eluents of different concentrations, concentrate and dry to obtain Berberis fruit extract B;

(3) The above-mentioned Berberis fruit extract A and Berberis fruit extract B, one or both of which are mixed in a certain proportion to obtain the Berberis fruit extract of the present invention.

The preparation method of barberry fruit extract is:

(1) Berberis fruit, water is used as solvent, the extraction temperature is 50°C-95°C, the number of extractions is 1-3 times, each extraction time is 1-4 hours, and the amount of each solvent is 6-15% of the weight of Berberis fruit times; filter, combine extracts, concentrate to relative density d=1.10-1.18, filter to obtain liquid A;

(2) Pass the medicinal liquid A obtained in step (1) through a non-polar macroporous adsorption resin, and first elute with water, and the water eluate directly passes through the polar macroporous adsorption resin, and then use a Elute the non-polar macroporous adsorption resin with ethanol solution, collect ethanol eluents with different concentrations, concentrate and dry to obtain Berberis fruit extract A; then elute the polar macroporous adsorption resin with ethanol solution with a concentration of 50%-95% Resin, collect ethanol eluents of different concentrations, concentrate and dry to obtain Berberis fruit extract B;

(3) Mix the above-mentioned Berberis fruit extract A and Berberis fruit extract B to obtain the Berberis fruit extract of the present invention.

The preparation method of barberry fruit extract is:

(1) Berberis fruit, use water as a solvent, reflux extraction, the number of extractions is 2 times, each extraction time is 2 hours, and the amount of solvent used for each time is 10 times the weight of Berberis fruit; filter, combine the extracts, and concentrate to Relative density d=1.12, filtered to obtain medicinal solution A;

(2) Pass the drug solution A obtained in step (1) through D101 non-polar macroporous adsorption resin, and first elute with water. , 60% ethanol solution to elute D101 non-polar macroporous adsorption resin sequentially, collect ethanol eluents of different concentrations, concentrate and dry to obtain Berberis fruit extract A; then use 70% and 95% ethanol solutions in sequence Elute the LSA-40 polar macroporous adsorption resin, collect ethanol eluents with different concentrations, concentrate and dry, and obtain Berberis fruit extract B;

(3) Mix the above-mentioned Berberis fruit extract A and Berberis fruit extract B to obtain the Berberis fruit extract of the present invention.

The berberberry fruit extract of the present invention mainly contains: berberine, jatrorrhizine, palmatine, magnolialine, geranium pigment, mallow pigment.

The preparation method of barberry fruit extract of the present invention is characterized in that: the non-polar macroporous adsorbent resin adopted is D101 macroporous adsorbent resin, AB-8 macroporous adsorbent resin; the polar macroporous adsorbent resin adopted is LSA- 40 macroporous adsorption resin, HPD600 macroporous adsorption resin.

The barberry fruit extract of the present invention is prepared into pharmaceutical oral dosage forms such as tablets, granules, and capsules by adding various pharmaceutically acceptable auxiliary materials.

Application of the barberry fruit extract A and the barberry fruit extract B of the present invention in the preparation of medicines for treating diabetes.

The medicine for treating diabetes composed of the barberry fruit extract of the invention and chemical medicine, traditional Chinese medicine or natural medicine.

The medicine for treating diabetes composed of berberis fruit extract A, berberberry fruit extract B and chemical medicine, traditional Chinese medicine or natural medicine of the present invention.

The present invention explores and researches for the first time that the dry and mature fruit of Berberis nummularia Bge. is used as a raw material to extract and prepare an extract for treating diabetes. Animal experiments have shown that barberry fruit extract has obvious hypoglycemic effect. The clinical application research of this experiment shows that Berberis fruit extract can significantly reduce urine protein and blood muscle intoxication in patients with DN, and improve renal function. The treatment group was better than the control group, but had no significant effect on fasting blood sugar (P>0.05). No obvious adverse reactions were found in the treatment group after treatment.

detailed description

The application of barberry fruit extract in the preparation of medicines for treating diabetes will be further described below through specific experimental examples and examples, but it is not limited to the present invention.

Embodiment 1: the preparation of berberis fruit extract and monomeric compound

(1) Berberis fruit 15kg, use water as solvent, reflux extraction, the number of extractions is 2 times, each extraction time is 2 hours, and the amount of solvent used for each time is 10 times the weight of Berberis fruit; filter, combine the extracts, concentrate To relative density d=1.12, filter to obtain medicinal liquid A;

(2) Pass the drug solution A obtained in step (1) through D101 non-polar macroporous adsorption resin, and first elute with water. , 60% ethanol solution to elute D101 non-polar macroporous adsorption resin sequentially, collect ethanol eluents of different concentrations, concentrate and dry to obtain Berberis fruit extract A; then use 70% and 95% ethanol solutions in sequence Elute the LSA-40 polar macroporous adsorption resin, collect ethanol eluents with different concentrations, concentrate and dry, and obtain Berberis fruit extract B;

(3) Mix the above-mentioned Berberis fruit extract A and Berberis fruit extract B to obtain the Berberis fruit extract of the present invention.

Silica gel column chromatography, preparative thin-layer chromatography, and SephadexLH-2O column chromatography were performed on the Berberis fruit extract to obtain ecdyberberine, jatrorrhizine, palmatine, magnolialine, geranium pigment, and mallow pigment, respectively. The chemical structures of the above compounds were confirmed by means of mass spectrometry and nuclear magnetic resonance, and the purity was greater than 98% by high performance liquid chromatography.

Embodiment 2: the preparation of berberis fruit extract and monomeric compound

(1) Berberis fruit 12kg, water is used as solvent, extraction temperature is 50°C, extraction times are 3 times, each extraction time is 1 hour, and the amount of solvent used for each time is 6 times the weight of Berberis fruit; filter and combine the extracts , concentrated to a relative density of d=1.18, filtered to obtain medicinal solution A;

(2) Pass the drug solution A obtained in step (1) through AB-8 non-polar macroporous adsorption resin, and first elute with water. ethanol solution to elute AB-8 non-polar macroporous adsorption resin, collect 50% ethanol eluate, concentrate and dry to obtain Berberis fruit extract A; then elute HPD600 polarity with 95% ethanol solution Macroporous adsorption resin, collecting 95% ethanol eluate, concentrating and drying to obtain Berberis fruit extract B;

(3) Mix the above-mentioned Berberis fruit extract A and Berberis fruit extract B to obtain the Berberis fruit extract of the present invention.

Silica gel column chromatography, preparative thin-layer chromatography, and SephadexLH-2O column chromatography were performed on the Berberis fruit extract to obtain ecdyberberine, jatrorrhizine, palmatine, magnolialine, geranium pigment, and mallow pigment, respectively. The chemical structures of the above compounds were confirmed by means of mass spectrometry and nuclear magnetic resonance, and the purity was greater than 98% by high performance liquid chromatography.

Embodiment 3: the preparation of berberis fruit extract and monomeric compound

(1) Berberis fruit 18kg, water is used as solvent, the extraction temperature is 95°C, the number of extractions is 1 time, each extraction time is 4 hours, and the amount of solvent used for each time is 15 times the weight of Berberis fruit; filter and combine the extracts , concentrated to a relative density of d=1.10, filtered to obtain medicinal solution A;

(2) Pass the drug solution A obtained in step (1) through D101 non-polar macroporous adsorption resin, and first elute with water, and the water eluate directly passes through HPD600 polar macroporous resin, and then use 95% ethanol solution Elute D101 non-polar macroporous adsorption resin, collect 95% ethanol eluate, concentrate and dry to obtain Berberis fruit extract A; then elute HPD600 polar macroporous adsorption resin with 50% ethanol solution, Collect 50% ethanol eluate, concentrate and dry to obtain Berberis fruit extract B;

(3) Mix the above-mentioned Berberis fruit extract A and Berberis fruit extract B to obtain the Berberis fruit extract of the present invention.

Silica gel column chromatography, preparative thin-layer chromatography, and SephadexLH-2O column chromatography were performed on the Berberis fruit extract to obtain ecdyberberine, jatrorrhizine, palmatine, magnolialine, geranium pigment, and mallow pigment, respectively. The chemical structures of the above compounds were confirmed by means of mass spectrometry and nuclear magnetic resonance, and the purity was greater than 98% by high performance liquid chromatography.

Embodiment 4: the preparation of barberry fruit extract tablet

Take 260g of the Berberis fruit extract of Example 1, add 55g of starch, mix evenly, granulate, dry, sieve, add 23g of microcrystalline cellulose, 3.5g of magnesium stearate, mix evenly, and compress into 1000 tablets, to obtain Berberry fruit extract tablet.

Embodiment 5: the preparation of barberry fruit extract capsule

Get Example 2 Berberis fruit extract 245g, add microcrystalline cellulose 10g, starch 40g, mix, granulate, dry, granulate, sieve, pack 1000 capsules, obtain the Berberis fruit extract capsule.

Embodiment 6: the preparation of barberry fruit extract granules

Get 265g of the Berberis fruit extract of Example 3, add 105g of dextrin, mix, granulate, dry, and granulate to obtain the Berberis fruit extract granules.

Experimental Example 1: Trial of berberis fruit extract on microangiopathy in type 2 diabetic nephropathy

Clinical data: A total of 40 patients were observed, including 22 males and 18 females, with an average age of 55 years. All patients had a history of diabetes for more than 10 years. All selected cases were in line with the diagnostic criteria for diabetes promulgated by WHO in 1999 and the diagnostic criteria for diabetic nephropathy in the sixth edition textbook "Internal Medicine" for colleges and universities. They were randomly divided into a treatment group and a control group of 20 cases each. There was no significant difference in gender, age, and disease course between the two groups (P>0.05), which were comparable. All patients had no acute or chronic nephritis and other kidney diseases, no urinary tract infection, fever and application of nephrotoxic drugs.

Treatment method: All patients were first given antihypertensive and hypoglycemic treatment, antihypertensive drugs used angiotensin-converting enzyme inhibitors (enalapril), hypoglycemic drugs used insulin (Ganshusen 30R), adjusted dosage according to the condition, so that fasting Blood sugar drops below 7mmol/L, blood pressure drops below 18.6/12.OkPa. The treatment group was additionally given tablets made of the barberry fruit extract of Example 4 of the present invention, 3 tablets/each time, 3 times a day, and 2 weeks was a course of treatment. The control group only used hypoglycemic, antihypertensive drugs and symptomatic treatment. Observe the changes of fasting blood glucose (FBG), 24-h urine protein quantification, blood-muscle compound clearance rate and endogenous muscle compound clearance rate in the two groups before and after treatment. Statistical processing: count data with X ² test, measurement data with t test.

Treatment results: the mean changes of FBG and 24h urine protein quantification, SCr and endogenous muscle clearance rate (CCr) in the two groups before and after treatment. After treatment, the 24-h urine protein quantification and blood muscle complex in the treatment group were significantly lower than those before treatment (P<0.01), and the clearance rate of endogenous muscle complex was significantly increased (P<0.01), and there was also a significant difference compared with the control group after treatment (P<0.01). <0.01). It shows that the addition of Berberis fruit extract treatment can significantly reduce urine protein and blood muscle intoxication in patients with DN, and improve renal function. The treatment group was better than the control group, but had no significant effect on fasting blood sugar (P>0.05). No obvious adverse reactions were found in the treatment group after treatment.

Experimental Example 2: Test of hypoglycemic effect of berberis fruit extract on spontaneously obese diabetic mice

Experimental animals: SPF grade, 50 spontaneously obese diabetic mice (db/db mice), 6 weeks old, weighing 38 ± 2g, were randomly divided into groups. Provided by the Animal Experiment Center of the Chinese Academy of Sciences.

Experimental drugs and instruments: Berberis fruit extract, Berberis fruit extract A, Berberis fruit extract B (prepared by the method in Example 1, batch numbers are: 20110405, 20110406, 20110407); the positive control drug is maleic acid Rosiglitazone (GlaxoSmithKline Tianjin Pharmaceutical Co., Ltd.), onetouchII blood glucose meter (Johnson & Johnson).

Grouping of animals: model control group: 10 animals, given the same amount of water; positive control group: 10 animals, given positive drug; experimental drug group: 30 animals, divided into three groups, respectively given Berberis fruit extract, Berberis fruit extract Extract A, Barberry Fruit Extract B.

Dosing: After being divided into groups, the dosing starts one week after adaptation, and the dosing lasts for 2 weeks. The dosing route is gavage. The doses of the three extracts converted to mice were 400 mg/kg body weight, and the dose of rosiglitazone was 5 mg/kg body weight.

Blood glucose measurement: 1 hour after administration on the 7th and 14th day, the blood was taken from the tail vein to measure the blood glucose of the animal immediately with a onetouch II blood glucose meter. The test results are shown in Table 1.

Table 1 Effects of barberry fruit extracts on blood glucose concentration in db/db mice

Note: *p<0.05, compared with model group.

The results in Table 1 show that each extract of Berberis fruit has a significant hypoglycemic effect on db/db mice. Among them, barberry fruit extract has the best hypoglycemic effect.

Claims (2)

1. the application of Berberis fruit extract in the preparation treatment diabetes medicine, the preparation method of described Berberis fruit extract is: (1) Berberis fruit, water is used as solvent, the extraction temperature is 50°C-95°C, the number of extractions is 1-3 times, each extraction time is 1-4 hours, and the amount of each solvent is 6-15% of the weight of Berberis fruit times; filter, combine extracts, concentrate to relative density d=1.10-1.18, filter to obtain liquid A; (2) Pass the medicinal liquid A obtained in step (1) through a non-polar macroporous adsorption resin, and first elute with water, and the water eluate directly passes through the polar macroporous adsorption resin, and then use a Elute the non-polar macroporous adsorption resin with ethanol solution, collect ethanol eluents with different concentrations, concentrate and dry to obtain Berberis fruit extract A; then elute the polar macroporous adsorption resin with ethanol solution with a concentration of 50%-95% Resin, collect ethanol eluents of different concentrations, concentrate and dry to obtain Berberis fruit extract B; (3) Mix the above-mentioned Berberis fruit extract A and Berberis fruit extract B to obtain the Berberis fruit extract. 2. application according to claim 1, is characterized in that the preparation method of described barberry fruit extract is: (1) Berberis fruit, use water as a solvent, reflux extraction, the number of extractions is 2 times, each extraction time is 2 hours, and the amount of solvent used for each time is 10 times the weight of Berberis fruit; filter, combine the extracts, and concentrate to The relative density is 1.12, filtered to obtain liquid A; (2) Pass the drug solution A obtained in step (1) through D101 non-polar macroporous adsorption resin, and first elute with water. , 60% ethanol solution to elute D101 non-polar macroporous adsorption resin sequentially, collect ethanol eluents of different concentrations, concentrate and dry to obtain Berberis fruit extract A; then use 70% and 95% ethanol solutions in sequence Elute the LSA-40 polar macroporous adsorption resin, collect ethanol eluents with different concentrations, concentrate and dry, and obtain Berberis fruit extract B; (3) Mix the above-mentioned Berberis fruit extract A and Berberis fruit extract B to obtain the Berberis fruit extract.