Application of compound polysaccharide in improvement of obesity type 2 diabetes mellitus

Abstract

The invention discloses a novel application of ginseng, poria cocos and atractylodes macrocephala koidz powder polysaccharide in improving obesity type 2 diabetes mellitus (T2DM), relates to a pharmacological action of the ginseng, poria cocos and atractylodes macrocephala koidz powder polysaccharide in improving obesity T2DM and application of the ginseng, poria cocos and atractylodes macrocephala koidz powder polysaccharide, in particular to preparation of the ginseng, poria cocos and atractylodes macrocephala koidz powder polysaccharide and application of the ginseng, poria cocos and atractylodes macrocephala koidz powder polysaccharide in medicines and health foods for treating obesity T2DM, and belongs to the technical field of traditional Chinese medicines. The ginseng, poria cocos and atractylodes macrocephala koidz powder polysaccharide is extracted, and pharmacological experiments prove that the ginseng, poria cocos and atractylodes macrocephala koidz powder polysaccharide has a relatively strong effect of improving obesity T2DM. The ginseng, poria cocos and atractylodes macrocephala koidz powder polysaccharide disclosed by the invention has relatively good activity in the aspect of improving the obesity T2DM function, small side effects, low price and relatively good druggability, and can be used for preparing medicines for improving the obesity T2DM function independently or in combination with other medicines. The compound can be prepared into granules, capsules, tablets and other dosage forms with pharmaceutically or food acceptable carriers or excipients.

Description

Technical Field

[0001] The present invention relates to the technical field of traditional Chinese medicine, and specifically to the application of a compound polysaccharide in improving obesity type 2 diabetes. The present invention relates to the pharmacological effects and applications of polysaccharides in the classic prescription Shenling Baizhu San, and specifically to the preparation of compound Shenling Baizhu San polysaccharides and the application of compound polysaccharides in Shenling Baizhu San as active ingredients in improving obesity type 2 diabetes. Background Art

[0002] Type 2 diabetes mellitus (T2DM) is a group of common metabolic endocrine diseases characterized by insulin resistance and / or impaired insulin secretion. In 2021, there were 529 million people with diabetes worldwide, of which 96.0% had type 2 diabetes (Global, regional, and national burden of diabetes from 1990 to 2021, with projections of prevalence to 2050: a systematic analysis for the Global Burden of Disease Study 2021[J]. Lancet, 2023, 402(10397): 203-234.). The occurrence and development of T2DM are closely related to obesity. The main pathogenesis of obese T2DM is insulin resistance and pancreatic β-cell damage (Luo Jintong, Zhang Lijing, Zhan Libin, et al. Effect of Shenling Baizhu San on pancreatic β-cell dedifferentiation in obese type 2 diabetic ZDF rats [J]. Chinese Journal of Traditional Chinese Medicine, 2023, 38(12):6034-6039.). Strong evidence consistently shows that obesity management can delay the progression of prediabetes to type 2 diabetes and has positive significance for the treatment of type 2 diabetes (American-Diabetes-Association-Professional Committee. 8. Obesity and Weight Management for the Prevention and Treatment of Type 2 Diabetes: Standards of Care in Diabetes-2024 [J]. Diabetes Care, 2024, 47(Suppl 1): S145-S157.). Therefore, it is of great significance and urgency to study the pathogenesis of obese T2DM and effective prevention and treatment methods.

[0003] The classic prescription Shenling Baizhu San comes from the Taiping Huimin Hejiju Fang of the Song Dynasty. It was included in the National Essential Medicines Catalogue in 2009 and is now included in the 2020 edition of the Pharmacopoeia of the People's Republic of China (Volume 1). The whole prescription is composed of 10 herbs: ginseng, Poria cocos, Atractylodes macrocephala, Chinese yam, white hyacinth bean, lotus seed, coix seed, Amomum villosum, Platycodon grandiflorum, and licorice. The whole prescription is mainly sweet (eight out of ten are sweet), and is a representative prescription for spleen deficiency and dampness (National Pharmacopoeia Committee. Pharmacopoeia of the People's Republic of China: 2020 Edition. Volume 1 [M]. Beijing: China Medical Science and Technology Press, 2020.). Moreover, there have been relevant records of the treatment of diabetes since ancient times, such as: "Zhengzhi Zhunsheng·Lianfang" (authored by Wang Kentang, annotated by Wu Wei et al. Zhengzhi Zhunsheng [M]. Beijing: China Traditional Chinese Medicine Press, 1997: 6.): "It is especially suitable for patients with diabetes"; "Zhengzhi Huibu·Xiaoke" (edited by Li Yongcui in the Qing Dynasty, compiled by Zhu Jianping. The must-read series of Chinese medicine clinical practice reprinted Zhengzhi Huibu [M]. Beijing: People's Medical Publishing House, 2023: 4.): "Therefore, Shenling Baizhu San is a magical medicine for curing the disease"; "Miscellaneous Diseases Guangyao·Xiaoke" (edited by Danbo Yuanjian. Miscellaneous Diseases Guangyao [M]. Beijing: People's Medical Publishing House, 1983: 3.): "When diabetes first occurs... after a long time, Shenling Baizhu San should be used"; "Yuanke Fahui·Spleen Diseases" (Ming Wanjin. Yuke Fahui [M]. Beijing: People's Medical Publishing House, 1959.): "It is used to treat spleen and stomach weakness, loss of appetite, fatigue and lack of energy...diabetes" and so on. Master of Traditional Chinese Medicine Zhou Zhongying pointed out in his clinical experience in diabetes that if the body fluid is deficient and cannot be transformed into qi, it will lead to deficiency of both qi and yin, and damage to both body fluid and qi... For those with weak spleen qi, use Shenling Baizhu San (written by Zhou Zhongying. Zhou Zhongying [M]. Beijing: China Medical Science and Technology Press, 2010.).

[0004] In modern clinical practice, Shenling Baizhu San is commonly used to treat digestive system diseases. In recent years, with the changes in the disease spectrum, it has been widely used in metabolic diseases such as obesity and type 2 diabetes according to the principle of treating different diseases with the same method, with definite efficacy (Jin Xiaoqin, Liu Xiaomin, Chen Hui, et al. Systematic evaluation of the effects of Shenling Baizhu San on blood sugar and related indicators in patients with type 2 diabetes [J]. World Chinese Medicine, 2023, 18(08): 1106-1114; Zeng Shaowu. Clinical efficacy observation of Shenling Baizhu San combined with metformin in the treatment of overweight / obese type 2 diabetic patients with spleen deficiency and dampness syndrome [D]. Hubei University of Chinese Medicine, 2022.). In basic research, Shenling Baizhu powder can reduce the body weight of obese type 2 diabetes (Zhang Xiaozhi. Effect of Shenling Baizhu powder on the body weight of obese type 2 diabetes [J]. Journal of Practical Chinese Medicine, 2010, 24(05):93-94.), improve obese T2DM by activating the AMPK signaling pathway and promote lipid and glucose metabolism, thereby alleviating lipid metabolism disorders (Zhang Shuqin, Li Yanjie, Qin Hewei, et al. Exploring the mechanism of action of Shenling Baizhu powder in improving obese type 2 diabetic mice based on AMPK signaling pathway [J]. Chinese Medicinal Materials, 2022, 45(11):2737-2742.), and can also improve the intestinal microbial imbalance of obese mice (Jin Xiaoqin, Liu Bin, Liu Haoyuan, et al. Exploring the mechanism of action of Shenling Baizhu powder in treating obesity based on intestinal flora [J]. Journal of Traditional Chinese Medicine, 2023, 38(08): 1732-1738.). Other studies have shown that Shenling Baizhu San can effectively improve glucose and lipid metabolism disorders and insulin resistance in rats with metabolic syndrome, reduce liver fatty degeneration and inflammatory cell infiltration, and thus alleviate the inflammatory state (Li Yi, Cai Huiling, Zhao Minhong, et al. Effects of Shenling Baizhu San on insulin resistance and inflammatory state in rats with metabolic syndrome [J]. Journal of Guangzhou University of Chinese Medicine, 2023, 40(08): 2026-2032.).This team took Professor Zhan Libin's theory of "spleen yin" as the guiding ideology, combined with previous work, and applied the classic prescription Shenling Baizhu San, which embodies the law of nourishing spleen yin with sweet and mild spleen, has the effects of strengthening spleen qi, nourishing spleen yin, and penetrating spleen dampness, and plays the role of ascending and descending, dispersing and collecting, and nourishing and moving at the same time. It is especially suitable for patients with spleen yin deficiency and dampness, that is, the basic prescription for the application of the theory of nourishing spleen yin (Luo Jintong, Zhang Lijing, Zhan Libin, et al. Effect of Shenling Baizhu San on pancreatic β-cell dedifferentiation in obese type 2 diabetic ZDF rats [J]. Chinese Journal of Traditional Chinese Medicine, 2023, 38(12):6034-6039; Hang Tianyi, Zhan Libin, Sun Tiefeng, et al. Effect of Shenling Baizhu San on sweet taste receptors in type 2 diabetic Zucker rats [J]. Chinese Journal of Traditional Chinese Medicine, 2023; Zhang Lijing, Zhan Libin, Hang Tianyi, et al. Study on the effect of Shenling Baizhu powder on improving chronic inflammation and preventing obesity and type 2 diabetes in ZDF rats based on intestinal flora[J]. Chinese Journal of Traditional Chinese Medicine, 2022, 47(04): 988-1000; Yan Yinghao. Study on the mechanism of action of Shenling Baizhu powder targeting intestinal G6pc to improve glucose and lipid metabolic diseases based on "spleen-intestine" correlation[D]. Nanjing University of Chinese Medicine, 2023; Zang Ningzi, Wang Shiwen, Li Pin, et al. Study on the development of spleen yin theory from ancient times to now and the interpretation of the effect of Shenling Baizhu powder on nourishing spleen yin based on literature and metrology methods[J]. Chinese Journal of Traditional Chinese Medicine, 2024, 42(03): 210-214; Zhang Lijing, Zhan Libin. Exploration of the material basis and mechanism of action of Shenling Baizhu powder in the treatment of type 2 diabetes based on integrative pharmacology platform[J]. Chinese Journal of Experimental Traditional Chinese Medicine, 2018, 24(21): 157-162.). Modern studies have found that most obese T2DM patients have a clear preference for fatty and sweet foods, so intervening in the sweet taste preference of obese T2DM patients is beneficial to the prevention and treatment of the disease.Obese T2DM patients show a preference for sweet taste (Min Gao, Jebb Susan-A, Aveyard Paul, et al. Associations Between Dietary Patterns and Incident Type 2 Diabetes: Prospective Cohort Study of 120,343 UK Biobank Participants[J]. Diabetes Care, 2022, 45(6):1315-1325.). This is because overeating fatty and sweet foods can hinder the spleen and produce dampness, affecting the production and distribution of fine substances, which indirectly reflects the body's lack of fine substances. Sweet drugs can strengthen the spleen and promote digestion. Therefore, a treatment strategy of "treating sweet with sweet" is proposed to prevent and treat obese T2DM caused by sweet taste preference, which is mainly based on sweet drugs, Shenling Baizhu San. It is also appropriate to follow the patient's preferences (Chen Yuan, Li Zhenhua, Yue Rensong, et al. On the application of sweet drugs in diabetes based on "following the appropriateness"[J]. Chinese Journal of Basic Medicine of Traditional Chinese Medicine, 2019, 25(12): 1739-1740.). However, compared with single herbs, Shenling Baizhu San as a traditional Chinese medicine compound is more complex, and its true anti-obesity type 2 diabetes drug efficacy material basis is still poorly understood, the effective components are not yet fully understood, and there are no reports on its effective parts (Lu Guangying, Xing Xunyan, Wang Jiayun, et al. Research progress of the classic prescription Shenling Baizhu San and prediction analysis of quality markers [J]. Chinese Journal of Traditional Chinese Medicine, 2022, 47(19): 5171-5181.). Related studies suggest that polysaccharide components may be the main components of Shenling Baizhu San in regulating intestinal flora and metabolites (Lin Ping. Study on the microecological mechanism of Shenling Baizhu San, a spleen-strengthening prescription, based on "traditional Chinese medicine polysaccharides-butyrate-producing bacteria" [D]. Jiangxi University of Traditional Chinese Medicine, 2021.). Therefore, the present invention explores the role of Shenling Baizhu San and its polysaccharides in intervening in obesity type 2 diabetes, and clarifies the main material basis of Shenling Baizhu San's "treating sweetness with sweetness".

[0005] The excavation and revelation of the effective ingredient group or effective ingredients in Shenling Baizhu San that play this role has positive significance for interpreting the compatibility changes and applications of prescriptions, and there is still a lack of systematic research. Through the search of literature books, no reports were found on the function of Lingbaizhu San polysaccharide in improving obesity T2DM. Summary of the invention

[0006] In view of the defects of the prior art, the purpose of the present invention is to provide a new use of polysaccharides from Shenling Baizhu San, that is, polysaccharides from Shenling Baizhu San have the function of improving obesity type 2 diabetes, can be used to prepare anti-sugar drugs and health products, and have a significant therapeutic effect on obesity type 2 diabetes. The research results show that polysaccharides are the main active ingredients of Shenling Baizhu San for improving obesity type 2 diabetes. Through pharmacological experiments, it is found that polysaccharides from Shenling Baizhu San show that they have a strong effect on improving obesity type 2 diabetes.

[0007] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions.

[0008] The invention discloses an application of a compound polysaccharide in preparing a product for improving obesity type 2 diabetes, characterized in that the compound polysaccharide is a Shenling Baizhu powder polysaccharide.

[0009] The present invention also discloses a method for preparing Shenling Baizhu powder polysaccharide, which is characterized by comprising the following steps: Step 1: Soak the medicinal materials in pure water 8 times the volume of the raw medicinal materials for 2 hours, heat and boil, then turn to low heat and distill for 0.5 hours, and filter with filter cloth to obtain the first decoction; add water 8 times the volume of the raw medicinal materials to the remaining medicinal residue, heat and boil, then turn to low heat and distill for 0.5 hours, and then filter with filter cloth to obtain the second decoction; Step 2: Combine the two decoctions obtained after distillation in step 1, and concentrate under reduced pressure until the volume of the solution is 1 times the volume of the raw medicinal material, slowly add anhydrous ethanol, adjust the ethanol concentration to 70%, stir while adding, let stand at 0-4°C overnight, and collect the precipitate; Step 3: pour out the supernatant, repeatedly sonicate the precipitate with anhydrous ethanol, and obtain the polysaccharide component after suction. After freeze-drying, the polysaccharide of Shenling Baizhu powder is obtained.

[0010] The present invention also discloses the use of the Shenling Baizhu powder polysaccharide prepared by the above-mentioned preparation method in the preparation of a drug for improving obesity type 2 diabetes.

[0011] The present invention also discloses a medicine for improving obesity type 2 diabetes, characterized in that the medicine contains the above-mentioned Shenling Baizhu San compound polysaccharide.

[0012] Furthermore, the dosage form of the compound polysaccharide of Shenling Baizhu San includes granules, capsules or tablets.

[0013] The present invention also discloses a pharmaceutical composition, characterized in that the composition comprises the above-mentioned Shenling Baizhu San compound polysaccharide and pharmaceutically acceptable excipients.

[0014] The invention also discloses the use of the above-mentioned pharmaceutical composition in preparing a drug for improving obesity and type 2 diabetes.

[0015] The present invention adopts a water extraction and alcohol precipitation method to prepare polysaccharides in Shenling Baizhu powder. Shenling Baizhu powder and its polysaccharides and liraglutide are used to treat obese T2DM rats. The body weight, abdominal circumference, food intake, water intake and fasting blood sugar of the rats are measured regularly, and oral glucose tolerance test (Oral glucose tolerance test, OGTT), insulin tolerance test (Insulintolerance test, ITT), pyruvate tolerance test (Pyruvate tolerance test, PTT) are performed after the administration. Parameters such as the homeostasis model-insulin resistance index (Homeostasis model assessment of insulin resistance index, HOMA-IR), the homeostasis model insulin secretion index (Homeostasismodel assessment of β-cell, HOMA-β), the insulin sensitivity index (Insulin sensitivity index, ISI), and the quantitative insulin sensitivity index (Quantitative insulin sensitivity check index, QUICKI) are calculated. At the end of the experiment, serum and pancreatic tissue are collected for biochemical and pathological analysis. The obese T2DM animal model was successfully established by using (Highfatdiet, HFD) + (Streptozotocin, STZ). Shenling Baizhu powder and its polysaccharides can reduce the body weight and abdominal circumference of obese T2DM rats, alleviate the phenotype of polydipsia and polyphagia in obese T2DM rats, and regulate lipid metabolism. It can reduce fasting blood sugar in obese T2DM rats, improve glucose tolerance, insulin tolerance and pyruvate tolerance, improve pancreatic islet morphology and function, and regulate sugar metabolism.

[0016] Compared with the prior art, the beneficial effects of the present invention are as follows.

[0017] 1. The present invention explores and discovers for the first time a new pharmacological effect of Shenling Baizhusan polysaccharide, namely, Shenling Baizhusan polysaccharide can resist obesity T2DM and improve obesity T2DM.

[0018] 2. The present invention is the first to apply Shenling Baizhu San polysaccharide to the preparation of drugs and health foods for improving the functions of obesity T2DM. It can be used alone or in combination with other drugs to treat obesity T2DM, and can also be made into granules, capsules, tablets and other dosage forms with pharmaceutically or food acceptable carriers or excipients. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1The effect of Shenling Baizhu powder and its polysaccharide on the body weight and abdominal circumference of obese T2DM rats. A: body weight of rats in each group from the beginning to the end of the experiment; B: body weight of rats in each group at the end of drug administration; C: abdominal circumference of rats in each group from the beginning to the end of the experiment; D: abdominal circumference of rats in each group at the end of drug administration; compared with group C, ### P <0.001,## P <0.01, # P <0.05; compared with group M,*** P <0.001,** P <0.01,* P <0.05.

[0020] Figure 2 The effect of Shenling Baizhu powder and its polysaccharide on the water intake of obese T2DM rats. A: Food intake of rats in each group from the beginning to the end of the experiment; B: Average food intake of each rat in each group at the end of drug administration; C: Water intake of rats in each group from the beginning to the end of the experiment; D: Average water intake of each rat in each group at the end of drug administration; Compared with group C, ### P <0.001,## P <0.01, # P <0.05; compared with group M,*** P <0.001,** P <0.01,* P <0.05.

[0021] Figure 3 The effects of Shenling Baizhu powder and its polysaccharides on glucose metabolism in obese T2DM rats. A: fasting blood glucose every week after administration; BD: FINS, GHbA1c, GLP-1; EF: comparison of OGTT values ​​and areas under the curve of rats in each group at the end of administration; GH: comparison of ITT values ​​and areas under the curve of rats in each group at the end of administration; IJ: comparison of PTT values ​​and areas under the curve of rats in each group at the end of administration; compared with group C, ### P <0.001,## P <0.01, # P <0.05; compared with group M,*** P <0.001,** P <0.01,* P <0.05.

[0022] Figure 4 The effect of Shenling Baizhu powder and its polysaccharide on lipid metabolism in obese T2DM rats. Compared with group C, ### P <0.001,## P <0.01, # P <0.05; compared with group M,*** P <0.001,** P<0.01,* P <0.05.

[0023] Figure 5 HOMA-IR, HOMA-β, ISI, QUICKI. Compared with group C, ### P <0.001,## P <0.01, # P <0.05; compared with group M,*** P <0.001,** P <0.01,* P <0.05.

[0024] Figure 6 HE staining of pancreatic tissue.

[0025] Figure 7 is the oxygen consumption rate of pancreatic homogenate. DETAILED DESCRIPTION

[0026] The present invention is further described in detail below with specific embodiments, but it should not be understood that the scope of the above subject matter of the present invention is limited to the following embodiments, and all technologies realized based on the content of the present invention belong to the scope of the present invention.

[0027] Unless otherwise specified, all reagents and materials used in the present invention are commercially available.

[0028] Example 1 Preparation of Shenling Baizhu powder polysaccharide.

[0029] The medicinal materials of Shenling Baizhu powder (each pack of Shenling Baizhu powder includes: 15 g ginseng, 15 g Poria, 15 g stir-fried Atractylodes, 15 g Chinese yam, 9 g lotus seeds, 9 g stir-fried Coix seeds, 12 g stir-fried white lentils, 6 g Amomum villosum, 6 g Platycodon grandiflorum, 9 g roasted Licorice root) were immersed in pure water 8 times the volume of the raw medicinal materials for 2 hours, heated and boiled, then turned to low heat and distilled for 0.5 hours, filtered with filter cloth to obtain the first decoction; the remaining medicinal residue was added with water 8 times the volume of the raw medicinal materials, heated and boiled, then turned to low heat and distilled for 0.5 hours, and then filtered with filter cloth to obtain the second decoction. The two decoctions obtained after distillation were combined and concentrated under reduced pressure until the volume of the solution was 1 times the volume of the raw medicinal materials, and anhydrous ethanol was slowly added thereto to adjust the ethanol concentration to 70%, stirred while adding, and allowed to stand overnight (0-4°C), and the precipitate was collected. The supernatant was poured out, and the precipitate was repeatedly sonicated with anhydrous ethanol, and the polysaccharide component was obtained after suction. After freeze-drying, the polysaccharide of Shenling Baizhu powder was obtained.

[0030] Example 2: Shenling Baizhu powder polysaccharide improves obesity and type 2 diabetes.

[0031] 1. Animal grouping and sample collection.

[0032] After one week of adaptive feeding, 9 rats were randomly selected from 45 healthy male SD rats as the blank control group (C) by the random number table method and fed with ordinary feed, while the remaining rats were fed with high-fat feed and had free access to food and water. Obese rats were obtained after feeding with a high-fat diet. All groups except the blank control group were injected intraperitoneally with a single rapid 1% STZ solution at a dose of 30 mg / kg body weight, while the blank control group was injected intraperitoneally with an equal amount of sodium citrate buffer. Further modeling was performed, and obese T2DM rats were obtained after random blood glucose screening. Obese T2DM rats (36 rats) were divided into the obese T2DM model group (M), the positive control group (liraglutide group, L), and the Shenling Baizhu powder ( Shenlingbaizhu Powder, SLBZP) and its polysaccharide (polysaccharidesfaction, PF) group. After modeling, the blank control group continued to be fed with ordinary feed, and the other groups were still fed with high-fat feed. The day after the successful modeling, oral gavage treatment was started, and 1 mL / 100 g body weight was administered for 4 weeks. According to the results of the optimal effective dose of Shenling Baizhu San decoction screened by the research team in the early stage (24 g kg -1 ), so the subsequent experiments will use this model to study the effect of Shenling Baizhusan decoction and its polysaccharides on obesity T2DM at this dose. According to the polysaccharide yield, it is converted into the corresponding multiple of crude drug dosage; the obese T2DM model group and the blank control group were given ultrapure water by gavage for 4 weeks in the same way.

[0033] At the end of the experiment, all animals were anesthetized with isoflurane after fasting for 12 h, blood was drawn through the abdominal aorta, and tissues were collected and stored in a -80°C refrigerator until detection.

[0034] 2.Indicator detection.

[0035] 2.1 General condition of rats: During the administration period, the body weight, abdominal circumference, body length, food intake and water intake of rats in each group were recorded every week.

[0036] 2.2 Lee's index: body weight (g)^(1 / 3)×10 / body length (mm), where body length is defined as the distance from nose to anus.

[0037] 2.3 Determination of fat body ratio: fat body ratio = (perinephric fat (g) + peritesticular fat (g)) / body weight (g) × 100%.

[0038] 2.4 Blood glucose testing: During the medication period, fasting blood glucose (FBG) was tested every week.

[0039] 2.5 OGTT, ITT, PTT.

[0040] OGTT, ITT, and PTT were performed at the end of the fourth week of treatment. OGTT: After fasting for 12 hours, rats were gavaged with 50% glucose solution at 2 g / kg body weight (converted to a solution volume of 0.4 mL / 100 g body weight), and blood glucose levels were measured by tail vein blood sampling before (0 min) and 30, 60, 90, and 120 min after taking sugar. ITT: After one day of adaptive feeding after OGTT, rats were fasted for 6 hours and intraperitoneally injected with regular short-acting insulin (0.5 U / kg body weight). Blood glucose levels were measured by tail vein blood sampling before (0 min) and 15, 30, 60, 90, and 120 min after injection. PTT: After one day of adaptive feeding after ITT, rats were fasted for 12 hours, weighed, and intraperitoneally injected with 2 g / kg of sodium pyruvate (prepared 2 g / 10 mL, 10 mL / kg), with an intraperitoneal injection volume of 1 mL / 100 g. Blood glucose levels were measured by sampling blood from the tail vein before injection (0 min) and 30, 60, 90, and 120 min after injection.

[0041] 2.6 Indicators related to sugar and lipid metabolism.

[0042] The levels of rat glycosylated hemoglobin A1c (HbA1c), rat fasting insulin (FINS), rat glucagon-like peptide-1 (GLP-1), and rat free fatty acid (FFA) were determined by enzyme-linked immunosorbent assay. The specific operations were carried out strictly in accordance with the instructions of the kit.

[0043] The levels of total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C) in serum were automatically measured using an automatic biochemical analyzer.

[0044] 2.7 Pancreatic islet morphology and functional indicators.

[0045] HOMA-IR=FBG×FINS / 22.5; HOMA-β=20×FINS / (FBG-3.5); ISI=1 / (FBG×FINS); QUICKI = 1 / (Log FBG+Log FINS); HE staining of pancreatic tissue.

[0046] 2.8 High-resolution respirometer (Oxygraph 2k, O2k) was used to measure the oxygen consumption rate of pancreatic tissue homogenate.

[0047] (1) Preparation of tissue homogenate: ① After taking the sample on ice, weigh it immediately, generally 5-10 mg / chamber (in vitro processing should be rapid), and place the remaining unweighed tissue in ice PBS for later use; ② Cut it into pieces; ③ Place the finely chopped tissue + 1 mL MIR05 in a manual glass homogenizer and homogenize until a suspension without fragments is obtained; ④ Filter out large residues after homogenization (if there are no large residues, filtration can be ignored), and the tissue homogenate is obtained. (2) O2k loading volume: 5-10 mg tissue + 1 mL MIR05 respiration medium per chamber, prepare 1 mL tissue homogenate. (3) Add sample and close the chamber: Remove the stopper, suck out 1 mL BUFFER in the chamber, use a pipette to add 1 mL tissue homogenate into the chamber, that is, the total volume in the chamber is 2 mL, and close the chamber for detection. Titration scheme: G+M+P+ADP(D)+CYTC+SCC(S)+U+ROT+AMA.

[0048] 2.9 Statistical methods.

[0049] The data were analyzed using Prsim 9.0 software, and the values ​​were expressed as mean ± standard deviation. One-way ANOVA was used to compare the means of multiple groups. P The difference was statistically significant when the value was <0.05.

[0050] 3 Experimental results.

[0051] 3.1 Effects of Shenling Baizhu San and its polysaccharides on body weight and abdominal circumference of obese T2DM rats.

[0052] The changes of body weight and abdominal circumference of rats in each group during the experiment were as follows: Figure 1 As shown. The body weight of rats fed with high-fat diet increased compared with the blank control group. After 4 weeks of high-fat diet feeding, the body weight of rats fed with high-fat diet showed an upward trend. After intraperitoneal injection of 1% STZ solution, the body weight of rats in group M, group L, SLBZP and its polysaccharide groups decreased. After administration, the body weight of rats in each group decreased compared with group M; at the 4th week of administration, group L ( P <0.001), SLBZP group ( P <0.01), PF group ( P <0.01) had a significant decrease in body weight.

[0053] After 4 weeks of high-fat diet feeding, the abdominal circumference of rats fed with high-fat diet was found to be higher than that of group C; after drug administration, the abdominal circumference of rats in each drug administration group tended to decrease compared with group M. P <0.0001), SLBZP group ( P <0.001), PF group (P <0.01) had a significant reduction in abdominal circumference.

[0054] Compared with group C, the Lee's index in group M tended to increase, and after administration, it tended to decrease, but there was no statistical significance ( P >0.05); compared with group C, the lipid body ratio of group M was significantly increased ( P <0.05), and there was a decreasing trend after administration.

[0055] 3.2 Effects of Shenling Baizhu Powder and its polysaccharides on water intake in obese T2DM rats.

[0056] The changes in food intake and water intake of rats in each group during the experiment were as follows: Figure 2 As shown. During the feeding process in the obesity stage, the rats fed with high-fat diet in the first 3 weeks ate less food than the blank control group of rats fed with ordinary diet, and the food intake was higher than that of group C in the 4th week. After intraperitoneal injection of 1% STZ solution, the food intake of rats in group M and each drug-treated group decreased before administration. In the absence of drug treatment, the food intake of group M increased compared with group C. After one week of administration, the food intake of rats in group M and each drug-treated group recovered. Starting from the second week of administration, the food intake of each drug-treated group decreased compared with group M ( P <0.01, P <0.001).

[0057] According to the results of water intake, in the first 4 weeks of high-fat diet feeding, the water intake of the blank control group showed an increasing trend compared with the rats fed with high-fat diet at the beginning, and the water intake of the rats fed with high-fat diet showed an increasing trend compared with the blank group in the next two weeks. After intraperitoneal injection of 1% STZ solution after the 4th week, the water intake of rats in groups M, L, SLBZP and its polysaccharide groups increased compared with that in group C, showing obvious characteristics of T2DM polydipsia. Compared with rats in group M, the water intake of rats in groups L, SLBZP and polysaccharide groups decreased significantly ( P <0.01).

[0058] 3.3 Effects of Shenling Baizhu San and its polysaccharides on glucose metabolism in obese T2DM rats.

[0059] 3.3.1 Effects of Shenling Baizhu powder and its polysaccharides on fasting blood glucose in obese T2DM rats.

[0060] Compared with group C, the FBG of T2DM rats injected with 1% STZ solution increased significantly. The FBG of T2DM rats showed that the FBG of all rats was greater than 11.1mM, and the T2DM modeling rate was good. With the administration of the drug, the drug group significantly reduced the FBG of obese T2DM rats compared with group M.

[0061] 3.3.2 Effects of Shenling Baizhu powder and its polysaccharides on routine biochemical indicators of glucose metabolism in obese T2DM rats.

[0062] At the end of the experiment, the expression of FINS, GHbA1c and GLP-1 in rats of each group was analyzed. The results showed that after the administration, the FINS of rats in group M increased significantly compared with that in group C ( P <0.001), compared with the M group, the FINS in each drug-treated group was significantly reduced, and the differences were statistically significant compared with the M group ( P <0.05).

[0063] 3.3.3 Effects of Shenling Baizhu powder and its polysaccharides on glucose tolerance, insulin tolerance and pyruvate tolerance in obese T2DM rats.

[0064] OGTT and ITT after one week of administration showed that compared with group C, the glucose regulation of obese T2DM rats was impaired to a certain extent, and glucose tolerance and insulin tolerance were abnormal. After one week of each drug intervention, the impaired glucose tolerance and insulin intolerance of T2DM rats had not been significantly improved. After the end of drug administration, the glucose tolerance, insulin tolerance and pyruvate tolerance of T2DM rats were improved.

[0065] 3.4 Effects of Shenling Baizhu Powder and its polysaccharides on lipid metabolism in obese T2DM rats.

[0066] Compared with group C, the levels of TC, TG, LDL-C, and FFA in the serum of rats in group M increased, while the level of HDL-C decreased. After the administration, compared with group M, the levels of TC, TG, LDL-C, and FFA in each administration group decreased, and the level of HDL-C increased. The TC, TG, and LDL-C levels of obese T2DM rats were improved, and the FFA content in the serum of rats was significantly reduced in group PF ( P <0.001), the PF group significantly increased the content of HDL-C in the serum of rats ( P <0.0001). These results indicate that SLBZP and its polysaccharides can improve lipid metabolism in obese T2DM rats.

[0067] 3.5 Effects of Shenling Baizhu powder and its polysaccharides on pancreatic islet morphology and functional indicators in obese T2DM rats.

[0068] 3.5.1 HOMA-IR, HOMA-β, ISI, QUICKI.

[0069] Compared with group C, HOMA-IR of rats in group M was significantly higher than that in group C ( P <0.0001), indicating that the established obese T2DM rat model had insulin resistance symptoms. After the drug intervention, the HOMA-IR of each drug-treated group decreased significantly, and the differences were statistically significant compared with the M group ( P<0.0001). Compared with group C, the HOMA-β index of rats in group M was significantly decreased ( P <0.0001). After the end of the drug intervention, the pancreatic β-cell function index of each drug group showed an upward trend, and the HOMA-β of the L group was significantly higher than that of the M group ( P <0.01). Compared with group C, the ISI of rats in group M was significantly reduced ( P <0.0001. After the drug intervention, the ISI of each medication group increased significantly ( P <0.001, P <0.0001). The QUICKI value of group M was significantly lower than that of group C ( P <0.0001), after the drug intervention, the QUICKI value increased significantly ( P <0.0001), indicating that Shenling Baizhu powder and its polysaccharides can significantly improve the insulin sensitivity of insulin-resistant rats.

[0070] 3.5.2 HE staining of pancreatic tissue.

[0071] Islet morphological changes Figure 6 As shown in the figure, the pancreatic islets of rats in group C were normal in morphology, oval in shape, with closely arranged islet cells and basically uniform nuclei. The pancreatic islets of rats in group M were severely damaged, with irregular shapes, irregular edges, uneven arrangement of islet cells, and changes in nuclei. The pancreatic islets of rats in the SLBZP and its polysaccharide groups were generally intact, mostly oval in shape, with relatively clear boundaries, and a few nuclei were compensatory enlarged, which was significantly improved compared with group M. The pancreatic islets of rats in group L were basically intact in morphology, with relatively uniform cell arrangement, compensatory enlargement of nuclei, and no obvious damage.

[0072] 3.5.3 O2k was used to measure the oxygen consumption rate of pancreatic homogenate.

[0073] Compared with group C, the oxygen consumption rate of pancreatic homogenate in group M was decreased; compared with group M, the oxygen consumption rate of pancreatic homogenate in groups L, SLBZP, and PF all tended to increase, indicating that Shenling Baizhu powder and its polysaccharides play an important role in pancreatic dysfunction and damage in obese T2DM.

[0074] The present invention prepares a large amount of Shenling Baizhu powder polysaccharide, and for the first time explores whether Shenling Baizhu powder polysaccharide has the function of improving obesity T2DM. After a large number of experimental studies, it is found that Shenling Baizhu powder polysaccharide has a good effect on improving obesity T2DM.

[0075] The above description is only a preferred embodiment of the present invention and is not intended to limit the scope of the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. An application of a compound polysaccharide in the preparation of a product for improving obesity and type 2 diabetes, characterized in that: The compound polysaccharide is Shenling Baizhusan polysaccharide.

2. A method for preparing Shenling Baizhu powder polysaccharide, characterized in that: The following steps are involved: Step 1: Soak the medicinal materials in pure water 8 times the volume of the raw medicinal materials for 2 hours, heat and boil, then turn to low heat and distill for 0.5 hours, and filter with filter cloth to obtain the first decoction; add water 8 times the volume of the raw medicinal materials to the remaining medicinal residue, heat and boil, then turn to low heat and distill for 0.5 hours, and then filter with filter cloth to obtain the second decoction; Step 2: Combine the two decoctions obtained after distillation in step 1, and concentrate under reduced pressure until the volume of the solution is 1 times the volume of the raw medicinal material, slowly add anhydrous ethanol, adjust the ethanol concentration to 70%, stir while adding, let stand at 0-4°C overnight, and collect the precipitate; Step 3: pour out the supernatant, repeatedly sonicate the precipitate with anhydrous ethanol, and obtain the polysaccharide component after suction. After freeze-drying, the polysaccharide of Shenling Baizhu powder is obtained.

3. Use of the Shenling Baizhusan polysaccharide prepared by the preparation method according to claim 2 in the preparation of drugs for improving obesity and type 2 diabetes.

4. A drug for improving obesity type 2 diabetes, characterized in that: The medicine contains the compound polysaccharide of Shenling Baizhu San according to claim 1.

5. The drug according to claim 3, characterized in that: The dosage form of the compound polysaccharide of Shenling Baizhu powder includes granules, capsules or tablets.

6. A pharmaceutical composition, characterized in that The composition comprises the compound polysaccharide of Shenling Baizhu San according to claim 1 and pharmaceutically acceptable excipients.

7. Use of the pharmaceutical composition according to claim 6 in the preparation of a drug for improving obesity and type 2 diabetes.

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