Glycine globulin compound for preventing hyperlipemia as well as preparation method and application of glycinin compound

Abstract

The invention discloses a glycinin compound for preventing hyperlipemia as well as a preparation method and application of the glycinin compound, and belongs to the field of biological medicines. The preparation method comprises the following steps: stirring and mixing resveratrol and a soybean 7S globulin solution at 40-60 DEG C for 1.5-2.5 h, so that resveratrol and soybean 7S globulin are fully combined to obtain a glycinin compound; the mass ratio of the soybean 7S globulin to the resveratrol is (250: 50)-(250: 45). Experimental results show that the glycinin compound disclosed by the invention can inhibit the weight increase of a hyperlipemia rat, remarkably reduce the fat index and the liver index, effectively improve the abnormal change of the blood fat factor level and show an obvious lipid-lowering effect. In-vitro simulated digestion experiments show that the glycinin compound shows peptide fragments with different strength from soybean 7S protein after gastrointestinal digestion, and meanwhile, the glycinin compound improves the in-vitro cholate binding capacity.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine, and particularly relates to a glycinin complex for preventing hyperlipidemia, a preparation method thereof, and an application thereof. Background Art

[0002] At present, most people take lipid-lowering drugs to control obesity and improve dyslipidemia. However, the side effects of these drugs are obvious. For example, orlistat can cause gastrointestinal reactions, pancreatic damage, and kidney damage. Statins can cause muscle damage, liver damage, and increase the incidence of diabetes, etc., and cannot meet the requirements of human health. Therefore, it is of great significance to develop new therapies that are safe, green, have positive functions but few side effects.

[0003] Plant extracts usually contain a variety of active ingredients and have the advantages of no side effects and low cost. In recent years, the research on plant extracts with lipid-lowering effects has become a hot topic. As a physiological function material, soy protein has been found to have a variety of physiological effects. A series of clinical studies have shown that consuming soy protein has a variety of beneficial health effects, such as reducing serum triglyceride and cholesterol levels, regulating blood pressure, and reducing risk factors for cardiovascular diseases. Soy 7S globulin is an active ingredient of soy protein and has been proven to improve the blood lipid status of animal models and human individuals, regulate lipid metabolism-related genes, and intestinal flora disorders. Therefore, soy 7S globulin has become a prospective functional food raw material for preventing and treating lipid metabolism-related diseases.

[0004] Polyphenols, as common plant extracts, have a variety of beneficial pharmacological effects, including anti-hyperlipidemia, anti-inflammatory, and antibacterial activities. Resveratrol is a natural polyphenol compound found in plants and has attracted great interest based on its protective effects in dyslipidemia and cardiovascular diseases. Resveratrol improves dyslipidemia by regulating key regulators of cholesterol and lipid metabolism (UCPI, PRDM16, and PGC-1α). Therefore, resveratrol is an effective drug for treating lipid metabolism-related diseases and has good prospects for clinical development.

[0005] Both soy 7S globulin and resveratrol are important components of food. In food processing, soy 7S globulin may combine with resveratrol to form a complex and coexist in a complex food system. Therefore, when soy 7S globulin and resveratrol are consumed in the form of a complex, it is not clear whether the soy globulin complex has a synergistic promoting effect on the regulation of blood lipid metabolism in hyperlipidemia patients, and it is of great significance for the development of prospective functional food raw materials for preventing and treating hyperlipidemia. Summary of the Invention

[0006] The present invention aims to address the deficiencies in the existing background technology, and provides a protein-polyphenol complex that is safe, green, has no side effects, and has the function of preventing hyperlipidemia, and discloses the application of the protein-polyphenol complex in lipid-lowering for patients with hyperlipidemia. Specifically, the present invention provides a glycinin complex for preventing hyperlipidemia, its preparation method and application.

[0007] To solve the above problems, the first aspect of the present invention is to provide a glycinin complex for preventing hyperlipidemia. Resveratrol and a 7S glycinin solution of soybean are stirred and mixed at 40 - 60 °C for 1.5 - 2.5 h to enable resveratrol to fully bind to the 7S glycinin of soybean, obtaining a glycinin complex (i.e., a 7S glycinin-resveratrol complex); the mass ratio of 7S glycinin of soybean to resveratrol is 250:50 - 250:45.

[0008] In a preferred embodiment, the 7S glycinin solution of soybean is a 1 - 10 wt% 7S glycinin solution of soybean.

[0009] Using the 7S glycinin solution of soybean within this concentration range enables the 7S glycinin of soybean to be uniformly dispersed in the solution, which helps the full binding of resveratrol to the 7S glycinin of soybean.

[0010] In a preferred embodiment, the 7S glycinin of soybean is prepared by the alkali solubilization and acid precipitation method.

[0011] In the glycinin complex, the binding of the 7S glycinin of soybean and the resveratrol complex is a non-covalent binding.

[0012] The second aspect of the present invention is to disclose a preparation method of a glycinin complex for preventing hyperlipidemia, including the following steps:

[0013] (1) Extraction of 7S glycinin of soybean: The defatted soybean powder is completely dissolved under alkaline conditions, and a 7S glycinin precipitate of soybean is obtained under acidic conditions;

[0014] (2) Preparation of the glycinin complex: The 7S glycinin of soybean is configured into a protein solution with a concentration of 1 - 10 wt% using deionized water, the pH is adjusted to 6.0 - 8.0, and it is stirred for 1.5 - 2.5 h; resveratrol is dispersed into the 7S glycinin solution of soybean so that the mass ratio of the 7S protein of soybean to resveratrol is 250:50 - 250:45, and then the mixture is stirred at 40 - 60 °C for 1.5 - 2.5 h to enable resveratrol to fully bind to the 7S glycinin of soybean; then, the free resveratrol that has not bound to the protein is removed, and it is freeze-dried to obtain a 7S-resveratrol complex.

[0015] The third aspect of the present invention is to provide the application of the glycinin complex in the preparation of a drug or food for preventing hyperlipidemia or in the preparation of a drug or food for enhancing the binding ability to bile salts.

[0016] In a preferred embodiment, hyperlipidemia refers to the increase in the contents of total cholesterol (TC), triglyceride (TG), and low-density lipoprotein (LDL-C) in the serum and the decrease in the content of high-density lipoprotein (HDL-C), and also includes hepatic steatosis, fat accumulation, decline in fat antioxidant level, liver injury, and intestinal flora disorder caused by hyperlipidemia.

[0017] Furthermore, the glycinin complex exhibits different degrees of digestion and different lengths of protein peptides compared with soybean 7S globulin after in vitro simulated gastrointestinal digestion, and the protein peptides obtained after in vitro simulated gastrointestinal digestion of the glycinin complex have a higher ability to bind bile salts.

[0018] The fourth aspect of the present invention is to disclose the application of the aforementioned glycinin complex in the preparation of a drug or food for controlling body weight, improving blood lipid factor levels, relieving liver injury, enhancing fat antioxidant levels, inhibiting fat accumulation, and promoting lipid metabolism, or in the preparation of a drug or food for regulating intestinal flora disorder, changing the intestinal flora structure, and increasing intestinal flora diversity.

[0019] The fifth aspect of the present invention is to provide a pharmaceutically or food-acceptable additive, which contains the aforementioned glycinin complex.

[0020] More specifically, soybean 7S globulin is prepared by the alkali dissolution and acid precipitation method, including: dissolving defatted soybean powder in deionized water, adjusting the pH to 8.0 - 10.0 with 1 - 2 mol / L NaOH solution, and stirring at room temperature for 0.5 - 2 h to obtain a slurry. Then, centrifuging the slurry to obtain a supernatant. Adding 0.05 - 0.1 mmol / L NaHSO3 to the supernatant, adding 5 - 8 mmol / L CaCl2, and adjusting the pH of the obtained solution to 6 - 6.4 with 2 mol / L hydrochloric acid. Keeping the supernatant at 0 - 4 °C overnight, then centrifuging and discarding the precipitate. Adjusting the pH of the centrifuged supernatant to 5.2 - 5.6 with 1 - 2 mol / L hydrochloric acid, and then centrifuging. Diluting the supernatant to twice its volume with ice water, adjusting the pH to 4.6 - 5.0, and collecting the precipitate after centrifugation as 7S. Before complete dissolution, washing the 7S fraction three times with deionized water and adjusting the pH to 7.2 - 7.8 with 1 - 2 mol / L NaOH solution. Dialyzing the obtained 7S fraction and then freeze-drying for standby.

[0021] Adding a reducing agent NaHSO3 with a concentration of 0.05 - 0.1 mmol / L and introducing 5 - 8 mmol / L of CaCl2 can increase the depolymerization between the two globulins of 11S and 7S, thereby improving the extraction rate of soybean 7S globulin.

[0022] More preferably, the defatted soybean powder is dissolved in deionized water at a mass - to - volume ratio of 1:15 - 1:10 (g / mL).

[0023] More preferably, during the preparation of soybean 7S globulin, the centrifugation speed is 6000 - 10000 rpm each time, and the centrifugation time is 20 - 30 min.

[0024] Preferably, the preparation of the soybean globulin complex includes the following steps: Dissolve a certain amount of soybean 7S globulin in deionized water to form a protein solution with a concentration of 2 wt%, then adjust the pH to 7.0 with 1 mol / L NaOH solution and stir for 2 h. Add an appropriate amount of resveratrol to the soybean 7S globulin solution, and then stir the mixture at 40 - 60 °C for 2 h to make resveratrol fully bind to soybean 7S globulin. Then, dialyze the complex at 4 °C to remove the free resveratrol that has not bound to the protein, and freeze - dry to obtain the soybean globulin complex.

[0025] More preferably, the mass ratio of soybean 7S globulin to resveratrol is 250:50 - 250:45.

[0026] More preferably, the stirring temperature of the mixture of soybean 7S globulin and resveratrol is 50 °C.

[0027] Adopting a slightly higher stirring and mixing temperature can accelerate the non - covalent binding of resveratrol and soybean 7S globulin to form a soybean globulin complex. However, the stirring and mixing temperature should not be too high to prevent protein denaturation.

[0028] The addition of resveratrol changes the digestion sites of protease in soybean 7S globulin, exposing different protein peptides, especially the protein peptides that are easy to regulate blood lipid metabolism, and improving the binding ability with bile salts.

[0029] The mass ratio of soybean 7S globulin to resveratrol is 250:50 - 250:45. Through the interaction between soybean 7S globulin and resveratrol, protein peptides that are more conducive to regulating blood lipid metabolism are exposed during digestion, thereby further improving the binding ability of soybean 7S globulin with bile salts and realizing the synergistic lipid - lowering effect of the soybean globulin complex. When the mass ratio of soybean 7S globulin to resveratrol is too high, the binding ability of the complex with bile salts is relatively weakened, and the peptides obtained after in vitro simulated digestion are relatively long; when the mass ratio of soybean 7S globulin to resveratrol is too low, the binding ability of the complex with bile salts is also relatively weakened.

[0030] In order to prove the effect of the soy globulin complex in preventing hyperlipidemia, the following methods were adopted:

[0031] (1) Determination of body weight, fat index and liver index;

[0032] (2) Determination of blood lipid factor levels;

[0033] (3) Detection of lipid accumulation by HE staining and Oil Red O staining;

[0034] (4) Evaluation of liver injury level;

[0035] (5) Determination of fat oxidative stress level;

[0036] (6) Detection of the expression of lipid metabolism-related genes by real-time quantitative PCR;

[0037] (7) Analysis of the structure and diversity of intestinal microflora;

[0038] (8) Analysis of peptide differences during in vitro gastrointestinal simulated digestion.

[0039] (9) Bile salt-binding ability after in vitro gastrointestinal simulated digestion.

[0040] Beneficial effects

[0041] 1. The present invention provides new ideas and methods for the prevention and treatment of hyperlipidemia and its complications. As natural plant extracts, soy 7S globulin and resveratrol are green, safe and have no toxic side effects. They can be administered by diet therapy, are easy to be accepted, and are conducive to clinical promotion and application.

[0042] 2. Resveratrol forms a complex with soy 7S globulin through interaction to protect it from being rapidly degraded in the gastrointestinal tract, thus significantly improving the functional effect of resveratrol in vivo. At the same time, the interaction between resveratrol and soy 7S globulin changes the digestion sites of proteases, exposes protein peptides that are easy to regulate lipid metabolism, and improves the binding ability with bile salts. This enables soy 7S globulin and resveratrol to have a synergistic promoting effect in regulating lipid metabolism and reducing cholesterol.

[0043] 3. The soy globulin complex of the present invention has a good improvement effect on the body weight, blood lipid levels in vivo and tissue lesion conditions of hyperlipidemic rats. The soy globulin complex can inhibit the increase in body weight and fat mass of rats fed a high-fat diet, reduce the levels of total cholesterol (TC), triglyceride (TG) and low-density lipoprotein (LDL-C) in the serum, increase the level of high-density lipoprotein (HDL-C); inhibit lipid accumulation and improve fat degeneration.

[0044] 4. The soy globulin complex of the present invention increases the level of superoxide dismutase (SOD) in the fat of hyperlipidemic rats, reduces the level of malondialdehyde (MDA) in the fat, and has good antioxidant activity on adipose tissue. At the same time, it also reduces the levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in the liver tissue and improves liver injury.

[0045] 5. The soy globulin complex of the present invention promotes the expression of the fat decomposition genes PRDM16, PGC-1α, and UCP1 in hyperlipidemic rats, indicating that the activation of the PRDM16 / PGC-1α / UCP1 pathway mediates the anti-hyperlipidemic effect of the soy globulin complex.

[0046] 6. The soy globulin complex of the present invention can regulate the intestinal flora disorder in hyperlipidemic rats, change the intestinal flora composition, and increase the α-diversity of the intestinal flora.

[0047] 7. The present invention reveals the improvement effect of the soy globulin complex on hyperlipidemia, which is expected to provide a new theoretical basis and new drug treatment targets for the treatment of hyperlipidemia and its complications, and provide a reference for the research of safe, green and non-toxic plant-derived pharmaceutical ingredients. Description of the Drawings

[0048] Figure 1 : Content diagrams of TC, TG, LDL-C, and HDL-C in the sera of rats in each group. A, TC content; B, TG content; C, LDL-C content; D, HDL-C content;

[0049] Figure 2 : SOD and MDA content diagrams in the white fat of rats in each group. A, SOD content; B, MDA content;

[0050] Figure 3 : AST and ALT content diagrams in the livers of rats in each group. A, AST content, B, ALT content;

[0051] Figure 4 : Stained diagrams of liver pathological tissue sections of rats in each group; A. HE staining diagram of the liver of rats in the normal control group; B. HE staining diagram of the liver of rats in the high-fat diet model group; C. HE staining diagram of the liver of rats in the soy 7S globulin experimental group; D. HE staining diagram of the liver of rats in the resveratrol experimental group; E. HE staining diagram of the liver of rats in the soy globulin complex experimental group; F. Oil red O staining diagram of the liver of rats in the normal control group; G. Oil red O staining diagram of the liver of rats in the high-fat diet model group; H. Oil red O staining diagram of the liver of rats in the soy 7S globulin experimental group; I. Oil red O staining diagram of the liver of rats in the resveratrol experimental group; J. Oil red O staining diagram of the liver of rats in the soy globulin complex experimental group;

[0052] Figure 5: Expression of UCP1, PRDM16, and PGC-1α genes in adipose tissue of rats in each group. A, UCP1; B, PRDM16; C, PGC-1α;

[0053] Figure 6 : Species abundance map of intestinal flora in rats in each group: A. Composition of intestinal flora at the phylum level; B. Composition of intestinal flora at the genus level; C. Alpha diversity - Chao index of intestinal flora; D. Alpha diversity - Shannon index of intestinal flora;

[0054] Figure 7 : SDS-PAGE map of protein complex after in vitro gastrointestinal simulated digestion. A, SDS-PAGE map after gastric digestion; B, SDS-PAGE map after intestinal digestion;

[0055] Figure 8 : Bile salt binding capacity of protein complex after in vitro gastrointestinal simulated digestion. A, Binding of Example 1 to sodium glycocholate; B, Binding of Example 1 to sodium taurocholate; C, Binding of Comparative Example 1 to sodium glycocholate; D, Binding of Comparative Example 1 to sodium taurocholate; E, Binding of Comparative Example 2 to sodium glycocholate; F, Binding of Comparative Example 2 to sodium taurocholate. Detailed implementation manners

[0056] The following combines the accompanying drawings and examples to illustrate the detailed implementation manners of the present invention. However, the following examples are only used to explain the present invention in detail and do not limit the scope of the present invention in any way.

[0057] Example 1:

[0058] (1) Extraction of soybean 7S globulin: Defatted soybean powder was stirred and dissolved with deionized water at a mass-to-volume ratio of 1:15 (g / mL), and the pH was adjusted to 9.0 using 2 mol / L NaOH solution. The mixture was stirred at room temperature for 1 h to obtain a slurry. Then, the slurry was centrifuged at 9000 rpm for 30 min to obtain the supernatant. Dry NaHSO3 was added to the supernatant (the concentration of NaHSO3 in the supernatant was 0.08 mmol / L), 5 mmol / L CaCl2 was further added, and the pH of the resulting solution was adjusted to 6.2 using 2 mol / L HCl. The supernatant was kept overnight at 4 °C, and then centrifuged at 6500 rpm for 20 min to discard the precipitate. The pH of the centrifuged supernatant was adjusted to 5.4 using 2 mol / L HCl, and then centrifuged at 9000 rpm for 30 min. The supernatant was diluted to twice its volume with ice water, the pH was adjusted to 4.8, and centrifuged at 6500 rpm for 20 min. The precipitate was collected as 7S. Before complete dissolution, the 7S fraction (i.e., the precipitate 7S) was washed three times with deionized water, and the pH of the washed 7S fraction solution was adjusted to 7.5 using 2 mol / L NaOH solution for complete dissolution. The final obtained 7S fraction solution was dialyzed at 3500 Da for 24 h and then freeze-dried for standby.

[0059] (2) Preparation of soybean globulin complex: 50 g of soybean 7S globulin was made into a protein solution with a concentration of 2 wt% using deionized water, and then the pH was adjusted to 7.0 using 1 mol / L NaOH and stirred at room temperature for 2 h. Resveratrol was dispersed into the soybean 7S globulin solution so that the mass ratio of soybean 7S globulin to resveratrol was 250:45. Subsequently, the mixture was stirred at 50 °C for 2 h to allow resveratrol to fully bind to soybean 7S globulin. Then, the complex was dialyzed at 8000 Da for 24 h to remove the free resveratrol that did not bind to the protein, and freeze-dried to obtain the soybean globulin complex.

[0060] Example 2:

[0061] Compared with Example 1 above, the difference lies in the extraction of soybean 7S globulin: Defatted soybean powder is stirred and dissolved with deionized water at a mass-to-volume ratio of 1:15 (g / mL), and the pH is adjusted to 9.0 using 2 mol / L NaOH solution, followed by stirring for 1 h at room temperature to obtain a slurry. Then, the slurry is centrifuged at 10,000 rpm for 20 min to obtain the supernatant. Dry NaHSO3 is added to the supernatant (the concentration of NaHSO3 in the supernatant is 0.08 mmol / L), 5 mmol / L CaCl2 is further added, and the pH of the resulting solution is adjusted to 6.2 using 2 mol / L HCl. The supernatant is kept overnight at 4 °C, and then centrifuged at 6,000 rpm for 30 min to discard the precipitate. The pH of the centrifuged supernatant is adjusted to 5.4 using 2 mol / L HCl, and then centrifuged at 10,000 rpm for 20 min. The supernatant is diluted to twice its volume with ice water, the pH is adjusted to 4.8, and centrifuged at 6,000 rpm for 30 min to collect the precipitate as 7S. Before complete dissolution, the 7S fraction (i.e., the precipitate 7S) is washed three times with deionized water, and the pH of the washed 7S fraction solution is adjusted to 7.5 using 2 mol / L NaOH solution for complete dissolution. The final obtained 7S fraction solution is dialyzed at 3,500 Da for 24 h and then freeze-dried for standby.

[0062] Example 3:

[0063] Compared with Example 1 above, the difference lies in the preparation of the soybean globulin complex: 50 g of soybean 7S globulin is formulated into a protein solution with a concentration of 2 wt% using deionized water, and then the pH is adjusted to 7.0 using 1 mol / L NaOH and stirred at room temperature for 2 h. Resveratrol is dispersed into the soybean 7S globulin solution such that the mass ratio of soybean 7S globulin to resveratrol is 250:50, and then the mixture is stirred at 50 °C for 2 h to allow resveratrol to fully bind to soybean 7S globulin. Then, the complex is dialyzed at 8,000 Da for 24 h to remove the free resveratrol that has not bound to the protein, and freeze-dried to obtain the soybean globulin complex.

[0064] Comparative Example 1:

[0065] Compared with Example 1 above, the difference lies in stirring the mixture at 25 °C for 2 h.

[0066] The specific steps are as follows:

[0067] (1) Extraction of soybean 7S globulin: Defatted soybean powder was stirred and dissolved in deionized water at a mass-to-volume ratio of 1:15 (g / mL), and the pH was adjusted to 9.0 using 2 mol / L NaOH solution. The mixture was stirred at room temperature for 1 h to obtain a slurry. Then, the slurry was centrifuged at 9000 rpm for 30 min to obtain the supernatant. Dry NaHSO3 was added to the supernatant (the concentration of NaHSO3 in the supernatant was 0.08 mmol / L), 5 mmol / L CaCl2 was further added, and the pH of the resulting solution was adjusted to 6.2 using 2 mol / L HCl. The supernatant was kept overnight at 4 °C, and then centrifuged at 6500 rpm for 20 min to discard the precipitate. The pH of the centrifuged supernatant was adjusted to 5.4 using 2 mol / L HCl, and then centrifuged at 9000 rpm for 30 min. The supernatant was diluted to twice its volume with ice water, the pH was adjusted to 4.8, and centrifuged at 6500 rpm for 20 min. The precipitate was collected as 7S. Before complete dissolution, the 7S fraction (i.e., the precipitate 7S) was washed three times with deionized water, and the pH of the washed 7S fraction solution was adjusted to 7.5 using 2 mol / L NaOH solution for complete dissolution. The final obtained 7S fraction solution was dialyzed at 3500 Da for 24 h, and then freeze-dried for standby.

[0068] (2) Preparation of soybean globulin complex: 50 g of soybean 7S globulin was prepared into a protein solution with a concentration of 2 wt% using deionized water, and then the pH was adjusted to 7.0 using 1 mol / L NaOH and stirred at room temperature for 2 h. Resveratrol was dispersed into the soybean 7S globulin solution so that the mass ratio of soybean 7S globulin to resveratrol was 250:45. Subsequently, the mixture was stirred at 25 °C for 2 h to allow resveratrol to fully bind to soybean 7S globulin. Then, the complex was dialyzed at 8000 Da for 24 h to remove the free resveratrol that did not bind to the protein, and freeze-dried to obtain Comparative Complex 1.

[0069] Comparative Example 2:

[0070] Compared with the above Example 1, the difference is that the mass ratio of soybean 7S globulin to resveratrol is 250:25.

[0071] The specific steps are as follows:

[0072] (1) Extraction of soybean 7S globulin: Defatted soybean powder was stirred and dissolved with deionized water at a mass-to-volume ratio of 1:15 (g / mL), and the pH was adjusted to 9.0 using 2 mol / L NaOH solution. It was stirred at room temperature for 1 h to obtain a slurry. Then, the slurry was centrifuged at 9000 rpm for 30 min to obtain the supernatant. Dry NaHSO3 was added to the supernatant (the concentration of NaHSO3 in the supernatant was 0.08 mmol / L), and 5 mmol / L CaCl2 was further added. The pH of the resulting solution was adjusted to 6.2 using 2 mol / L HCl. The supernatant was kept overnight at 4 °C, and then centrifuged at 6500 rpm for 20 min to discard the precipitate. The pH of the centrifuged supernatant was adjusted to 5.4 using 2 mol / L HCl, and then centrifuged at 9000 rpm for 30 min. The supernatant was diluted to twice its volume with ice water, and the pH was adjusted to 4.8. It was centrifuged at 6500 rpm for 20 min, and the precipitate was collected as 7S. Before complete dissolution, the 7S fraction (i.e., the precipitate 7S) was washed three times with deionized water. The pH of the washed 7S fraction solution was adjusted to 7.5 using 2 mol / L NaOH solution for complete dissolution. The final obtained 7S fraction solution was dialyzed at 3500 Da for 24 h, and then freeze-dried for standby.

[0073] (2) Preparation of soybean globulin complex: 50 g of soybean 7S globulin was prepared into a protein solution with a concentration of 2 wt% using deionized water, and then the pH was adjusted to 7.0 using 1 mol / L NaOH and stirred at room temperature for 2 h. Resveratrol was dispersed into the soybean 7S globulin solution so that the mass ratio of soybean 7S globulin to resveratrol was 250:25. Subsequently, the mixture was stirred at 50 °C for 2 h to enable full binding of resveratrol to soybean 7S globulin. Then, the complex was dialyzed at 8000 Da for 24 h to remove the free resveratrol that was not bound to the protein, and freeze-dried to obtain the comparative complex 2.

[0074] Example 4: Intervention test of soybean globulin complex on hyperlipidemic rats

[0075] The soybean globulin complex involved below all refers to the soybean globulin complex prepared by the method of Example 1.

[0076] 1) Animal grouping and drug administration

[0077] Male Sprague-Dawley rats (4 weeks old, weighing 100 - 120 g), provided by Beijing Vital River Laboratory Animal Technology Co., Ltd., were fed in an SPF-level environment with controlled temperature (23 ± 1°C) and humidity (55 ± 10%), and a 12-hour light / dark cycle. After one week of adaptation, the rats were randomly divided into 5 groups of 6 each, namely: ① Normal diet control group (LFD): fed normal feed; ② High-fat diet model group (HFD): fed high-fat feed (45% fat for energy, 1.2% cholesterol, 0.2% sodium cholate); ③ High-fat diet - soybean 7S globulin experimental group (7S): fed high-fat feed and intragastrically administered soybean 7S globulin at a dose of 250 mg / kg / d. The soybean 7S globulin used in the 7S group was prepared by the method of step (1) of Example 1; ④ High-fat diet - resveratrol experimental group (RES): fed high-fat feed and intragastrically administered resveratrol at a dose of 45 mg / kg / d; ⑤ High-fat diet - soybean globulin complex experimental group (7S-RES): fed high-fat feed and intragastrically administered the soybean globulin complex prepared in Example 1 at a dose of 250 mg / kg / d. The LFD and HFD groups were intragastrically administered normal saline as a control. All rats had free access to food and water, and their body weights were measured once a week.

[0078] 2) Collection and preservation of biological samples

[0079] All rats were fed for 7 weeks, euthanized after an overnight fast, blood samples were collected, left to clot at room temperature for 30 min, centrifuged at 3000 rpm for 10 min at 4°C, the serum was collected and stored at -80°C. Tissue samples such as brown fat, white fat, and liver of the rats were quickly isolated, rinsed with normal saline, blotted dry with filter paper, weighed, aliquoted, snap-frozen in liquid nitrogen and stored at -80°C. Another part of the liver and white fat tissues were fixed in 10% formalin for subsequent staining, sectioning, and histopathological observation. Fresh fecal samples of the rats were collected at the end of the experiment and stored at -80°C.

[0080] 3) Detection of physiological and biochemical indexes

[0081] The changes in body weight, fat index, and liver index of rats in each group were calculated, and the levels of total cholesterol (TC), triglyceride (TG), high-density lipoprotein (HDL-C), low-density lipoprotein (LDL-C), superoxide dismutase (SOD), malondialdehyde (MDA), aspartate aminotransferase (AST), and alanine aminotransferase (ALT) in the serum of rats in each group were detected according to the instructions of the corresponding kits (Nanjing Jiancheng Bioengineering Institute).

[0082] Fat index (%) = fat weight / rat body weight

[0083] Liver index (%) = Liver weight / Body weight of rats

[0084] 4) Histopathological analysis

[0085] The fixed liver tissues were paraffin-embedded, sectioned (about 5 μm), and dehydrated with gradient alcohol. Histological observations were performed using an optical microscope after hematoxylin-eosin (HE) and Oil Red O staining.

[0086] 5) RT-qPCR analysis

[0087] (1) RNA extraction and concentration determination: According to the experimental operation regulations, total RNA was extracted from inguinal white adipose tissue (IngWAT) using Trizol. Subsequently, 1 μL of the extracted total RNA sample was used to measure the RNA absorbance on a ultra-micro nucleic acid and protein analyzer (BioDrop, Cambridge, UK). The concentration of total RNA was diluted to about 1000 ng / μL with DEPC water and stored for further analysis. According to the instruction manual, cDNA was synthesized using a reverse transcription kit (Trans Gen). The cDNA was stored at -20 °C in a refrigerator for further analysis.

[0088] (2) Real-time fluorescence quantitative PCR: Real-time fluorescence quantitative PCR analysis was performed on a PCR instrument (T100, BIO-RAD, USA). The reaction system was as follows: 2.5 μL of cDNA, 1 μL of each upstream and downstream primer, 12.5 μL of 2×qPCR Mix, and 8.0 μL of ddH2O. The reaction conditions were as follows: pre-denaturation at 95 °C for 10 min, denaturation at 95 °C for 15 s, annealing at 60 °C for 1 min, and extension at 72 °C for 2 min, for 40 cycles. The relative mRNA expression levels of target genes were calculated using the 2 -ΔΔCT method, and β-actin was used as an internal reference gene for normalization control. The primer sequences used are shown in Table 1. The primer sequences in Table 1 are SEQ ID NO.1-8 from top to bottom:

[0089] Table 1 Primer sequences

[0090]

[0091] 6) Analysis of intestinal microbiota structure and diversity

[0092] According to the manufacturer's protocol, total DNA was extracted from the samples using the Fast DNA extraction kit (MP Biomedicals, CA, USA). The V3-V4 hypervariable region of the 16S rRNA gene was amplified by PCR using the forward primer 5'-ACTCCTACGGGAGGCAGCAG-3' and the reverse primer 5'-GGACTACHVGGGTWTCTAAT-3' (SEQ ID NO. 9-10). The polymerase chain amplification of the 16S rDNA domain was performed using the 16S rDNA primers for the V3-V4 region. High-throughput sequencing performed by Sangon Biotech (Shanghai) Co., Ltd. was used to determine the DNA sequences of the samples. All sequences were demultiplexed and quality-filtered using QIIME (version 1.9.1), and then data analysis was carried out.

[0093] 7) In vitro simulated digestion

[0094] Gastric digestion: A gastric juice medium was prepared by mixing 0.62 g of NaCl, 0.22 g of KCl, 50 mg of gastric lipase, 0.03 g of CaCl2·2H2O, 47.2 g of pepsin, and 0.12 g of NaHCO3 to make 200 ml, and adjusting the pH value to 3.0. 5 g of the sample (the sample was the soy 7S globulin or resveratrol obtained in step (1) of Example 1 or the soy globulin complex obtained in Example 1, Comparative Examples 1 and 2) was mixed with 500 mL of the artificial gastric juice medium and incubated at 37°C. Samples were taken at 0, 1 h, and 6 h respectively, and boiled in a 70°C water bath for 10 min to inactivate the enzymes.

[0095] Intestinal digestion: A intestinal juice medium was prepared by mixing 0.216 g of NaCl, 0.026 g of KCl, 0.0132 g of CaCl2·2H2O, and 5.6 mg of trypsin to make 40 ml, and mixing it with 40 ml of 7% pancreatin solution and 80 ml of 4% bile salt solution, and adjusting the pH value to 7.0. The intestinal juice was mixed with the liquid after gastric digestion at a ratio of 10:3 (v / v), and the pH value was adjusted to 7.0. Incubation was carried out at 37°C, and samples were taken at 0, 1, 2, and 3 h respectively, and boiled in a 70°C water bath for 10 min to inactivate the enzymes.

[0096] 8) SDS-PAGE

[0097] Mix 100 μL of the prepared soy 7S globulin and soy globulin-resveratrol complex sample (here, the sample refers to the sample taken during in vitro digestion in 7) with the loading buffer (33 μL), and add 10 μL of the obtained mixture to a 5% stacking gel and a 10% separating gel for electrophoresis. After staining with Coomassie Brilliant Blue for 3.5 h, decolorize with 5 w / v% methanol and 7.5 w / v% acetic acid. The gel image was obtained using a Bio-Rad imaging system (Bio-Rad Laboratories, Hercules, USA).

[0098] 9) Ability to bind bile salts in vitro

[0099] Prepare standard solutions of sodium glycocholate and sodium taurocholate at 10 mg / L and dilute them proportionally. Prepare 60% sulfuric acid. Add the sample solution and 60% sulfuric acid in a ratio of 1:4 (v / v) and mix. First, perform a water bath and then an ice bath. Measure the absorbance at 387 nm and plot the standard curves of sodium glycocholate and sodium taurocholate.

[0100] 10) Statistical analysis

[0101] Perform statistical analysis using SPSS 20.0 and GraphPad Prism 5.0. The data results are expressed as the mean ± standard deviation (X ± SD). One-way analysis of variance is used for processing, and the LSD test is used for pairwise comparison. P < 0.05 is considered to be of significant significance.

[0102] 11) Experimental results

[0103] I. The effects of the soy globulin complex on the body weight, fat index, and liver index of hyperlipidemic rats are shown in Table 2. After 7 weeks of feeding, compared with the control group, the body weight, brown fat index, white fat index, and liver index of the model group rats were significantly increased; compared with the model group, the body weight, brown fat index, white fat index, and liver index of the soy 7S globulin experimental group and the resveratrol experimental group rats were slightly decreased, but the differences were not significant. However, the brown fat index and white fat index of the soy globulin complex experimental group rats were significantly decreased, and the degrees of decrease in body weight and liver index were also better than those of the soy 7S globulin experimental group and the resveratrol experimental group alone. This indicates that a high-fat diet can cause an increase in rat body weight, fat accumulation, and a certain degree of fatty liver in the liver, while the intervention of soy 7S globulin and resveratrol can reduce the abnormal increase in body weight of hyperlipidemic rats, reduce fat accumulation, and simultaneously inhibit the formation of fatty liver. It should be noted that the intervention effect of the soy globulin complex is better than that of the individual protein and individual polyphenol.

[0104] Table 2 Changes in body weight, fat index, and liver index of rats in each group

[0105]

[0106] II. Effects of soy globulin complex on blood lipid factors in hyperlipidemic rats: Figure 1 As shown, compared with the control group, the levels of TC, TG, and LDL-C in the serum of rats in the model group were significantly increased, but the level of HDL-C was significantly decreased; compared with the model group, the TC levels in the 7S soy globulin experimental group and the resveratrol experimental group decreased by 43.9% and 37.3% respectively, the TG levels decreased by 21.2% and 22.8% respectively, the level of LDL-C decreased but not significantly, and the level of HDL-C increased but not significantly. However, the levels of TC, TG, and LDL-C in the soy globulin complex experimental group of rats decreased significantly, by 51.7%, 39.5%, and 34.8% respectively, and the level of HDL-C increased significantly by 49.0%, and the level was closer to that of the control group, with no significant difference. This indicates that the soy globulin complex can more effectively inhibit the increase of TC, TG, and LDL-C and the decrease of HDL-C in hyperlipidemic rats, and significantly improve the abnormal changes in the blood lipid levels of hyperlipidemic rats.

[0107] III. Effects of soy globulin complex on the levels of SOD, MDA in adipose tissue and AST, ALT in liver of hyperlipidemic rats:

[0108] Superoxide dismutase (SOD) can reduce tissue oxidative stress and play an antioxidant role; malondialdehyde (MDA) is a product of lipid peroxidation in organisms, which can cause cross-linking polymerization of proteins, nucleic acids, etc. and has cytotoxicity. As Figure 2 shown, compared with the control group, the level of SOD in the adipose tissue of rats in the model group showed a significant downward trend, and the level of MDA showed a significant upward trend; compared with the model group, the SOD levels in the 7S soy globulin experimental group and the resveratrol experimental group increased by 8.3% and 8.4% respectively, and the level of MDA hardly changed. However, the SOD level in the soy globulin complex experimental group increased significantly by 13.6%, and the MDA level decreased significantly by 13.0%. This shows that the soy globulin complex can better relieve the oxidative stress response of adipose tissue in hyperlipidemic rats and has good antioxidant activity. Aspartate aminotransferase (AST) and alanine aminotransferase (ALT) are important indicators reflecting liver function and can judge the degree of liver damage. As Figure 3As shown, compared with the control group, the levels of AST and ALT in the livers of rats in the model group were significantly increased, indicating impaired liver function. Compared with the model group, the levels of AST and ALT in the livers of rats in the 7S globulin experimental group, resveratrol experimental group, and globulin complex experimental group were significantly decreased. Among them, the levels of AST and ALT in the 7S globulin experimental group decreased by 24.5% and 25.6% respectively; the levels of AST and ALT in the resveratrol experimental group decreased by 24.7% and 18.9% respectively. It is worth noting that the degrees of decrease in AST and ALT in the globulin complex experimental group were greater, decreasing by 36.6% and 33.0% respectively, almost approaching those of the control group. This indicates that the globulin complex can better improve the liver function of hyperlipidemic rats and relieve liver damage.

[0109] IV. Effects of globulin complex on tissue lesions in hyperlipidemic rats

[0110] The liver is the main site of fat synthesis. As Figure 4 shown, the results of HE staining showed that the hepatic tissue structure of rats in the control group was intact, the hepatic lobule structure was clear, the hepatocytes were arranged neatly, and the cytoplasm showed a normal morphological structure. In contrast, the hepatic tissue structure of rats in the model group was loose, the hepatic lobule structure was unclear, the hepatocytes were swollen and increased in volume, and a large number of lipid vacuoles of different sizes appeared in the cytoplasm, indicating obvious fatty degeneration. Compared with the model group, the hepatic tissue and hepatic lobule structures in the 7S globulin experimental group and resveratrol experimental group became slightly clearer, the volume of hepatocytes decreased slightly, and the lipid vacuoles decreased slightly. However, in the globulin complex experimental group, the hepatic tissue and hepatic lobule structures became significantly clearer, the volume of hepatocytes decreased, the lipid vacuoles decreased significantly, and the overall morphology of the liver was closer to that of the normal group. The results of Oil Red O staining showed that almost no red lipid droplets were observed in the hepatocytes of rats in the control group; while a large number of obvious red lipid droplets were observed in the hepatocytes of rats in the model group, indicating very serious liver fat accumulation. Compared with the model group, the area and depth of red lipid droplets in the hepatocytes of rats in the 7S globulin experimental group and resveratrol experimental group were slightly reduced. In the globulin complex experimental group, the area and depth of red lipid droplets in the hepatocytes of rats were significantly reduced. The above results indicate that individual 7S globulin and individual resveratrol can play a certain preventive role in liver fatty degeneration, while the globulin complex can significantly improve liver fatty degeneration, reduce fat accumulation, and simultaneously show an obvious lipid-lowering effect.

[0111] V. Effects of globulin complex on the expressions of adipose UCP1, PGC-1α, and PRDM16 in hyperlipidemic rats

[0112] UCPI can generate heat by burning fatty acids and glucose, preventing abnormal fat accumulation; PGC-1α can regulate lipid metabolism and induce adipocyte differentiation, and has become a new target for treating diseases such as obesity; PRDM16 reduces the inflammatory response of the high-sugar and high-fat environment to the human body by promoting adipocyte browning and regulating adipokine levels. As Figure 5 shown, compared with the control group, the expression of UCP1, PGC-1α and PRDM16 genes in the model group rats was significantly down-regulated; compared with the model group, the expression of UCP1, PGC-1α and PRDM16 genes in the soy 7S globulin experimental group and the resveratrol experimental group rats was slightly up-regulated, but there was no significant difference. However, the expression of UCP1, PGC-1α and PRDM16 genes in the soy globulin complex experimental group rats was significantly up-regulated, tending to the control group. This indicates that the soy globulin complex can promote lipid metabolism, promote fat browning and regulate blood lipid factor levels. At the same time, it was confirmed that the activation of the UCP1 / PGC-1α / PRDM16 pathway mediated the anti-hyperlipidemic effect of the soy globulin complex.

[0113] VI. Effects of soy globulin complex on the intestinal flora structure and diversity of hyperlipidemic rats

[0114] The intestinal flora is an important internal environmental factor in the pathogenesis of hyperlipidemia and its related metabolic syndromes, and is regarded as a potential therapeutic target for such diseases. A high-fat diet will disrupt the structural composition and diversity of the body's intestinal flora. As Figure 6As shown in A-B in [reference], at the phylum level, the intestinal flora of rats is mainly composed of Firmicutes and Bacteroidetes. Compared with the control group, the abundances of Firmicutes and Bacteroidetes in the model group of rats decreased; compared with the model group, the abundances of Firmicutes and Bacteroidetes in the experimental groups of soybean 7S globulin, resveratrol, and soybean globulin complex all showed an upward trend. At the genus level, compared with the model group, the relative abundances of Akkermansia, Bacteroides, and Ruminococcus_torques_group, NK4A214_group, Ruminococcus_gnavus_group in rats that ingested the soybean globulin complex in the experimental group of soybean globulin complex increased, while the relative abundances of norank_Desulfovibrionaceae, Lachnospiraceae_NK4A136_group, norank_Lachnospiraceae, unclassified_Lachnospiraceae, Lactobacillus, Romboutsia, and unclassified_Oscillospiraceae decreased.

[0115] The α-diversity of the intestinal flora can reflect the richness and evenness of the flora, and the Chao index and Shannon index reflect species richness. As Figure 6 shown in C-D in [reference], compared with the control group, the Chao index and Shannon index of the intestinal flora of rats in the model group showed a downward trend, with no significant difference in the Chao index; compared with the model group, the Chao index and Shannon index of the intestinal flora of rats in each experimental group increased. This indicates that soybean 7S globulin, resveratrol, and soybean globulin complex can all regulate the intestinal flora structure and increase the diversity of the intestinal flora to a certain extent.

[0116] VII. Changes in Peptides in Soybean Globulin Complex after In Vitro Simulated Digestion

[0117] SDS-PAGE was used to identify the changes in peptides of protein digestion products and preliminarily identify the effect of resveratrol on the digestion peptides of soybean 7S globulin. As Figure 7As shown, there were no significant differences in the electrophoretic bands of undigested soy 7S globulin and soy glycinin complexes. After gastric digestion, the intensities of the large molecular weight bands (100 - 180 kDa, 60 - 72 kDa, and 30 - 50 kDa) of the soy glycinin complex decreased significantly, while the intensities of the small molecular weight bands (11 - 17 kDa and 22 - 25 kDa) increased significantly, indicating that long peptide segments were broken down into short peptide segments by proteases. Compared with soy 7S globulin, the bands near 11 - 17 kDa and 22 - 25 kDa of the soy glycinin complex after gastric digestion were darker in color, indicating that polyphenols enhanced the degree of protein digestion. After intestinal digestion, the obtained bands were mainly distributed around 25 kDa, with the top band disappearing and no large molecular weight bands observed. After intestinal digestion, the bands of the soy glycinin complex were closer to the bottom and had a smaller molecular weight than those of soy 7S globulin. This preliminarily demonstrated that the interaction between resveratrol and soy 7S globulin changed the digestion sites of proteases, exposing different protein peptide segments.

[0118] VIII. Bile salt - binding ability of soy glycinin complex after in vitro simulated digestion

[0119] When proteins bind to bile salts, they can effectively prevent the reabsorption of bile acids, promote defecation in the body. At the same time, in order to maintain the dynamic balance of bile acid fluid in the body, the liver will continuously convert cholesterol into bile salts, reduce the loss of bile salts, thereby reducing the cholesterol content in the body, and ultimately reducing the cholesterol content in the blood. By measuring the bile salt - binding ability in vitro, the lipid - lowering ability of the soy 7S protein - resveratrol complex can be evaluated to verify its lipid - lowering activity. As Figure 8 shown, compared with soy 7S protein alone and resveratrol alone, the bile salt (sodium glycocholate and sodium taurocholate) - binding ability of the soy 7S protein - resveratrol complex was stronger after gastrointestinal digestion. Calculate the average bile salt - binding rates at 0, 1, 2 h of gastric digestion and 1, 2, 3 h of intestinal digestion. Among them, the bile salt - binding ability of the soy 7S protein - resveratrol complex with sodium glycocholate and sodium taurocholate was increased by 16.0% and 14.0% respectively compared with that of soy 7S protein alone with sodium glycocholate and sodium taurocholate; the bile salt - binding ability of the soy 7S protein - resveratrol complex with sodium glycocholate and sodium taurocholate was increased by 9.0% and 8.6% respectively compared with that of resveratrol alone with sodium glycocholate and sodium taurocholate. Combining with the SDS - PAGE results, it can be shown that the interaction between resveratrol and proteins exposes protein peptide segments that are easy to regulate lipid metabolism.

[0120] After gastrointestinal digestion, the binding abilities of the soybean 7S protein-resveratrol complex in Comparative Example 1 with sodium glycocholate and taurocholic acid decreased by 10.2% and 7.4% respectively compared with those in Example 1; the binding abilities of the soybean 7S protein-resveratrol complex in Comparative Example 2 with sodium glycocholate and taurocholic acid decreased by 14.2% and 8.6% respectively compared with those in Example 1. In summary, the bile salt binding rates of the comparative complexes 1 and 2 prepared in Comparative Examples 1 and 2 were overall lower than that of the complex prepared in Example 1, indicating that the soybean 7S protein-resveratrol complex needs to be prepared under suitable conditions to most effectively bind to bile salts and exert its lipid-lowering function.

[0121] The experimental results of the soy globulin complex of the present invention show that it can inhibit the weight gain of hyperlipidemic rats, significantly reduce the fat index and liver index, effectively improve the abnormal changes in blood lipid factor levels, including significantly reducing TC, TG and LDL-C, increasing HDL-C, improving hepatic steatosis, reducing fat accumulation, enhancing the antioxidant activity of fat, alleviating liver injury, and promoting lipid metabolism by activating the UCP1 / PGC-1α / PRDM16 signaling pathway, showing an obvious lipid-lowering effect. In addition, it can also regulate the intestinal flora disorder caused by hyperlipidemia, change the intestinal flora structure, and increase the intestinal flora diversity. The in vitro simulated digestion experiment shows that the soy globulin complex shows peptide segments with different intensities from those of soybean 7S protein after gastrointestinal digestion, and at the same time, the soy globulin complex improves the in vitro bile salt binding ability.

[0122] Although the present invention has been disclosed above with preferred embodiments, it is not intended to limit the present invention. Anyone familiar with this technology can make various modifications and decorations without departing from the spirit and scope of the present invention. Therefore, the protection scope of the present invention should be defined by the claims.

Claims

1. A glycinin complex for preventing hyperlipidemia, characterized in that resveratrol and a 7S glycinin solution of soybean are stirred and mixed at 40 - 60 °C for 1.5 - 2.5 h to fully combine resveratrol with 7S glycinin of soybean, thereby obtaining a glycinin complex; the mass ratio of 7S glycinin of soybean to resveratrol is 250:50 - 250:

45.

2. The glycinin complex for preventing hyperlipidemia according to claim 1, wherein The 7S glycinin solution of soybean is a 1 - 10 wt% 7S glycinin solution of soybean.

3. The glycinin complex for preventing hyperlipidemia according to claim 1, wherein The 7S glycinin of soybean is prepared by an alkali - dissolution and acid - precipitation method.

4. The glycinin complex for preventing hyperlipidemia according to claim 1, characterized in that, In the glycinin complex, the combination of 7S glycinin of soybean and the resveratrol complex is a non - covalent bond.

5. A preparation method of a glycinin complex for preventing hyperlipidemia, characterized in that, It includes the following steps: (1) Extraction of 7S glycinin of soybean: The defatted soybean powder is completely dissolved under alkaline conditions, and a 7S glycinin precipitate of soybean is obtained under acidic conditions; (2) Preparation of the glycinin complex: The 7S glycinin of soybean is configured into a protein solution with a concentration of 1 - 10 wt% using deionized water, the pH is adjusted to 6.0 - 8.0, and it is stirred for 1.5 - 2.5 h; resveratrol is dispersed into the 7S glycinin solution of soybean so that the mass ratio of 7S protein of soybean to resveratrol is 250:50 - 250:45, and then the mixture is stirred at 40 - 60 °C for 1.5 - 2.5 h to fully combine resveratrol with 7S glycinin of soybean; then, the free resveratrol not bound to the protein is removed, and it is freeze - dried to obtain a 7S - resveratrol complex.

6. Use of the glycinin complex according to any one of claims 1 - 4 in the preparation of a drug or food for preventing hyperlipidemia or in the preparation of a drug or food for increasing the binding ability with bile salts.

7. The application according to claim 6, wherein Hyperlipidemia refers to the increase in the content of total cholesterol (TC), triglyceride (TG), and low - density lipoprotein (LDL - C) in serum and the decrease in the content of high - density lipoprotein (HDL - C) in serum. At the same time, it also includes hepatic steatosis, fat accumulation, decreased fat antioxidant level, liver damage, and intestinal flora disorder caused by hyperlipidemia.

8. The application according to claim 6, wherein The glycinin complex shows different digestion degrees and different lengths of protein peptides from 7S glycinin of soybean after in vitro simulated gastrointestinal digestion, and the protein peptides obtained after the glycinin complex undergoes in vitro simulated gastrointestinal digestion have a higher ability to bind bile salts.

9. Use of the glycinin complex according to any one of claims 1 - 4 in the preparation of a drug or food for controlling body weight, improving blood lipid factor levels, relieving liver damage, increasing fat antioxidant levels, inhibiting fat accumulation, and promoting lipid metabolism, or in the preparation of a drug or food for regulating intestinal flora disorder, changing the intestinal flora structure, and increasing intestinal flora diversity.

10. A pharmaceutically or food acceptable additive, characterized in that: The additive contains the glycinin complex according to any one of claims 1 - 4.

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