Application of black corn polysaccharide in preparing drugs for treating diseases related to intestinal barrier injury

Abstract

The present invention provides an application of black corn polysaccharide in the preparation of a drug for treating diseases related to intestinal barrier damage. By investigating its intestinal protective effect on a dextran sulfate sodium-induced mouse intestinal injury model, the research results show that black corn polysaccharide can improve clinical symptoms such as loose stools, weight loss, and colon atrophy caused by intestinal injury in mice, significantly reduce the disease score, inhibit the necrosis of goblet cells and mucosal edema and exfoliation in colon tissues, and enhance the intestinal mucosal barrier function. As a natural food commonly favored by the public, black corn has high safety and is expected to become an effective source of active ingredients for anti-intestinal injury, and has broad development prospects in protecting the intestinal barrier.

Description

Technical Field

[0001] The present invention belongs to the field of natural medicine chemistry, and particularly relates to the application of black corn polysaccharide in the preparation of drugs for treating diseases related to intestinal barrier damage. Background Art

[0002] The intestine is the main medium between humans and the environment. The intestine is not only responsible for absorbing essential dietary nutrients, but also for protecting the host from various ingested toxins and microorganisms, and intestinal barrier integrity is one of the important signs of intestinal health. The intestinal barrier is composed of normal intestinal flora, mucus layer, soluble antibacterial molecules, intestinal epithelial cell layer and intestinal immune system. Among them, the intestinal mucus barrier (mucus secreted by goblet cells) is the first barrier to prevent microorganisms from contacting the intestinal epithelium and plays an important role in maintaining intestinal barrier function. As the largest barrier tissue, the integrity of the intestinal barrier is indispensable for the body to maintain normal physiological functions. When the integrity of the intestinal barrier is damaged and barrier injury occurs, microorganisms and endotoxins in the intestine can break through the intestinal mucosal barrier, enter the blood, cause bacterial and endotoxin translocation, and promote the occurrence of intestinal origin infection, and even develop into systemic inflammatory response syndrome or multiple organ failure. A large number of studies have shown that the damage of the intestinal barrier is associated with the occurrence, development and metastasis of many diseases through the "brain-gut axis", "liver-gut axis", etc., including gastrointestinal diseases such as inflammatory bowel disease and colon cancer, as well as diabetes, non-alcoholic fatty liver, obesity, and even hyperglycemia, stroke, etc. Protecting the intestinal barrier can ensure the stability of the body's internal environment and reduce the occurrence of various diseases. Therefore, developing drugs to protect the intestinal barrier for treating related diseases has become a hot topic in clinical practice.

[0003] The more commonly used methods for improving intestinal barrier function include probiotic therapy and drug therapy. Probiotic therapy includes fecal microbiota transplantation or probiotic preparations, which maintain the balance of the microecosystem and protect the normal functions of the body through complex interaction reactions with the host and indigenous flora. However, probiotic treatment has certain side effects, such as easy to cause flatulence, abdominal distension, constipation and headache in patients. At the same time, long-term use of artificially synthesized probiotic products will gradually cause the intestine to lose its own function of reproducing probiotics, and over time, it is easy to develop dependence on probiotics. At present, there are still some drugs for improving intestinal barrier function, but most of them have poor effects and often strong side effects. Natural products have received more and more attention due to their mildness and safety. However, there are few natural functional foods for protecting the intestinal barrier and promoting intestinal health at present, and there is an urgent need to develop related products.

[0004] Black corn (Black corn, Zea mays L.), also known as black waxy corn and purple corn, belongs to the class Monocotyledoneae of the Angiospermae and is a plant of the genus Zea in the order Poales. Black corn is rich in nutrients, with higher contents of protein, amino acids, fat, and selenium than yellow corn, and contains a large amount of functional melanin and essential trace elements for the human body such as Zn, Fe, Ca, and P. Selenium is an essential trace element for the human body and plays an important role in many physiological processes. The selenium content in black corn can reach 3 to 8.5 times that of ordinary corn. Black corn also contains abundant polysaccharide substances. Studies have shown that polysaccharides are substances with biological activity. They can activate immune cells, improve the immune function of the body, and some also have antiviral and anti-aging effects, showing great potential in the health food market. However, there are few studies on the efficacy of polysaccharides in black corn kernels.

[0005] Black corn is easy to grow, has multiple yields in one season, and its production areas are spread all over the country. Such a huge production volume can ensure a large source of raw materials for the development of the black corn-related industry. However, in the current new trend of the combination of medicine and food and the joint use of medicine and food, the development of the black corn-related industry lags behind. There is an urgent need to conduct in-depth research on the natural extracts of black corn to accelerate the development of the black corn-related industry. Therefore, this invention studies the protective effect of black corn kernel polysaccharides on intestinal barrier injury, aiming to obtain a drug composition or functional product with the ability to protect the intestinal barrier and / or improve intestinal barrier injury. Summary of the Invention

[0006] This invention investigated the intestinal barrier protection activity of black corn polysaccharide solutions at different concentrations through in vivo activity experiments.

[0007] An intestinal injury animal model was established with dextran sulfate sodium, and different doses of black corn polysaccharide solutions were administered by gavage. The experimental results showed that black corn polysaccharide had a good therapeutic effect on intestinal injury caused by dextran sulfate sodium, and the high-dose effect was better, showing a certain dose-dependence. Specifically: (1) Black corn polysaccharide could improve the clinical symptoms of loose stools, weight loss, and colon atrophy caused by intestinal injury in mice, and reduce the disease score; (2) Black corn polysaccharide could inhibit the necrosis of goblet cells and mucosal edema and exfoliation in the colon tissue, and enhance the intestinal mucosal barrier function. The above research results indicate that black corn polysaccharide can play an important role in the preparation of preparations for protecting the intestinal barrier. Brief Description of the Drawings

[0008] Figure 1 Results of the effect of black corn polysaccharide on body weight change;

[0009] Figure 2 Results of the effect of black corn polysaccharide on disease score;

[0010] Figure 3 Results of the effect of black corn polysaccharide on colon length;

[0011] Figure 4 It is the macroscopic picture of mouse colon;

[0012] Figure 5 It is the result of H&E staining of mouse colon tissue. Specific implementation mode

[0013] To make the purpose, technical solutions and advantages of the present invention clearer, the following will further describe the implementation modes of the present invention in detail. However, the implementation modes of the present invention are not limited thereto. For process parameters or conditions not specifically noted, conventional techniques can be referred to.

[0014] Example 1 Preparation of black corn polysaccharide

[0015] (1) Separate black corn kernels from the cob and perform drying treatment; soak the dried corn kernels in 95% ethanol at a volume ratio of 1:8 overnight, repeat three times, then dry in the sun, and then crush and sieve to obtain black corn powder;

[0016] (2) Weigh 500 g of dry corn flour and put it into a beaker. Add distilled water according to the solid-liquid ratio of 1:5, and perform ultrasonic extraction at 60 °C for 1 hour each time, for a total of 4 times to obtain an extract; combine the extracts, filter to obtain a supernatant; perform rotary evaporation on the supernatant until the volume of the supernatant is reduced to 1 / 10 to obtain a concentrated solution;

[0017] (3) Add absolute ethanol to the concentrated solution until the ethanol content in the concentrated solution is 80%, place it in a refrigerator at 4 °C for 24 h, and centrifuge to obtain a crude polysaccharide precipitate;

[0018] (4) Finally, through concentration and freeze-drying, black corn crude polysaccharide is obtained, with an extraction rate of 48%, a polysaccharide content of 82.65%, a flavonoid content of 0.06%, and a selenium content of 0.3675 μg / g.

[0019] Example 2 Experiment on the protection of intestinal barrier by black corn polysaccharide

[0020] 1. Grouping and treatment of experimental animals

[0021] Experimental animals: 42 SPF-grade C57BL / 6 mice, male, 8 weeks old, with a body weight of 22 ± 2 g, provided by the Guangdong Provincial Laboratory Animal Center;

[0022] Experimental grouping: Randomly divided into 6 groups, including a normal group, a model group, a positive drug mesalazine group (200 mg / kg / d), a low-dose black corn polysaccharide group (400 mg / kg / d), a medium-dose black corn polysaccharide group (600 mg / kg / d), and a high-dose black corn polysaccharide group (800 mg / kg / d).

[0023] 2. Preparation of intestinal injury mouse model

[0024] After the mice were adaptively fed for 1 week, the model group and the treatment group began to establish the model and intervene in the treatment. Weigh 3 g of dextran sulfate sodium and dissolve it in 100 mL of distilled water for the mice in the model group and the treatment group to drink freely, and give the mice in the normal group distilled water to drink. The mice were given 3% dextran sulfate sodium solution for 7 days to induce an intestinal injury model. At the same time, the mice in each dose group of black corn polysaccharide and the mesalazine group were intragastrically administered the corresponding dose of the drug, and the mice in the normal group and the model group were intragastrically administered an equal volume of normal saline.

[0025] 3. Preventive and therapeutic effects of black corn polysaccharide on dextran sulfate sodium-induced intestinal injury in mice

[0026] During the whole experiment, the body weight, defecation and rectal bleeding of the mice were observed and recorded every day, and the disease activity score was calculated according to Table 1. On the 8th day, blood was collected from the orbital sinus, and then the mice were sacrificed. The length of the colon from the proximal end of the rectum to the ileocecal part was measured, and the distal colon tissue was immediately fixed with 4% paraformaldehyde solution or stored at -80 °C for further analysis.

[0027] For pathological tissue analysis of the colon tissue, part of the colon was fixed with 4% paraformaldehyde, embedded in paraffin, sectioned and stained with hematoxylin and eosin to evaluate the severity of intestinal injury.

[0028] Table 1 Disease activity scoring criteria

[0029]

[0030] 4. Experimental results and analysis

[0031] 4.1 Effects of black corn polysaccharide on body weight change, disease activity score and colon length

[0032] As Figures 1 to 4 shown, by the 8th day of the experiment, the average body weight of the mice in the normal group increased by 7.8%, the average disease activity score was 0, the colon was visible with complete feces, and there was no bloody stool; the body weight of the mice in the model group decreased by 13.8%, the average disease activity score was 10, the colon length was significantly atrophied, there was no complete feces in the colon, the colon was red and swollen and had obvious bloody stools; the body weight of the mice in the high-dose group of black corn polysaccharide decreased by 7.2%, the average disease activity score was 7.7, the degree of colon atrophy decreased, and there was less bloody stool in the colon. The above results indicate that the black corn polysaccharide of the present invention has a good protective effect on the body weight loss and colon atrophy symptoms caused by intestinal injury.

[0033] 4.2 Effects of black corn polysaccharide on intestinal barrier

[0034] As Figure 5As shown, compared with the normal group, the histological structure of the colon in the model group animals changed significantly. Goblet cells were severely necrotic and missing, the colonic mucosa layer exfoliated, the cell gaps were incomplete, there were irregular glandular structures, and the colonic mucosal barrier was severely damaged. After treatment with black corn polysaccharide, the histological structure of the colon in mice was significantly improved, the crypts were relatively intact, the number of goblet cells was significantly higher than that in the model group, the cells were more plump, and the intestinal mucosal barrier was relatively intact.

[0035] The above results indicate that the black corn polysaccharide prepared according to the present invention can enhance the function of the intestinal mucosal barrier, inhibit colonic mucosal edema, and inhibit the occurrence and development of dextran sulfate sodium-induced intestinal injury.

[0036] The difference between the present invention and the prior art lies in:

[0037] (1) Extracted substance: The prior literature reports the application of black corn ethanol extract. The black corn polysaccharide prepared in this patent is obtained by water extraction and ethanol precipitation combined with a series of refining means. Different extraction methods result in different extracted substances. At the same time, the prior literature only reports the application of corn silk polysaccharide extract. The black corn polysaccharide prepared in this patent is extracted from black corn kernels. Different sources lead to different structural compositions of the extracts. Moreover, the extraction rate of corn silk polysaccharide reported in the literature is often 16 - 40%, while the extraction rate of black corn kernel polysaccharide obtained by the method of this patent reaches 48%. The present invention discovers for the first time that black corn polysaccharide has the function of protecting the intestinal barrier, which can greatly expand the scope of use of black corn.

[0038] (2) Experimental model: This patent uses a mouse intestinal injury model induced by feeding dextran sulfate sodium to investigate indicators such as mouse body weight change, colon length, and degree of colonic tissue injury, so as to evaluate the protective effect of black corn polysaccharide on the intestinal barrier. This efficacy is studied for black corn for the first time, which is different from other literature.

[0039] The above is the preferred implementation manner of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and refinements can be made, and these improvements and refinements should also be regarded as the protection scope of the present invention.

Claims

1. Use of black corn polysaccharide in the preparation of a drug for treating intestinal injury, wherein: The preparation method of the black corn polysaccharide is as follows: (1) After drying the black corn kernels, soak them in 95% ethanol at a volume ratio of 1:8 overnight, dry them in the sun and then crush and sieve them to obtain black corn powder; (2) Add distilled water to the black corn powder at a material-liquid ratio of 1:5, and extract it by ultrasonic wave at 60 °C to obtain an extract; after filtering the extract, obtain the supernatant; perform rotary evaporation on the supernatant until the volume of the supernatant is reduced to 1 / 10 to obtain a concentrated solution; (3) Add anhydrous ethanol to the concentrated solution until the final concentration of ethanol is 80%, place it in a refrigerator at 4 °C for 24 h, and centrifuge to obtain a crude polysaccharide precipitate; (4) Finally, obtain it through concentration and freeze-drying.

2. The application according to claim 1, characterized in that The symptoms of the intestinal injury include at least one of loose stools, weight loss, colon atrophy, necrosis of goblet cells in the colon tissue, and mucosal edema and exfoliation.

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