A feed additive for improving intestinal health and preventing and treating intestinal diseases of animals and application thereof

Abstract

The application discloses a feed additive for improving intestinal health of animals and preventing and treating intestinal diseases and application thereof, and relates to the technical field of green and healthy breeding. The application firstly confirms that indole-3-butyric acid (IBA) has no toxic effect on cells and can relieve oxidative stress damage and inflammation damage of the cells in an effective concentration. Meanwhile, the application also fills the application blank of IBA as one of indole derivatives in livestock production, and provides experimental basis and theoretical basis for the fact that IBA is expected to be a new green additive for replacing antibiotics and zinc oxide. More importantly, the application also finds that IBA has positive effects in aspects of resisting diarrhea, repairing intestinal barrier damage, relieving intestinal inflammation and preventing and treating intestinal related diseases.

Description

Technical Field

[0001] This invention belongs to the field of green and healthy aquaculture technology, specifically involving the application of indole-3-butyric acid in the preparation of feed additives for improving animal intestinal health and preventing intestinal diseases. Background Technology

[0002] Weaning is a crucial step in the rearing and management of piglets. However, in actual production, after weaning, piglets rely entirely on feed for nutrition. Furthermore, leaving their sows and needing to be transferred to new groups and environments can cause weaning stress in piglets. Weaning stress negatively impacts the morphology and function of the piglet's small intestine, reducing digestive and absorptive capacity, thereby damaging the intestinal barrier function. This is accompanied by lethargy, ultimately leading to reduced feed intake, increased diarrhea rates, and stunted growth. In severe cases, it can even cause illness and death in piglets.

[0003] Adding a certain concentration of antibiotics to feed is a traditional practice to improve the growth performance of piglets and alleviate diarrhea rates. However, long-term use of antibiotics can lead to the emergence of drug-resistant bacteria, and antibiotic residues not only cause environmental pollution but also raise concerns about the quality and safety of livestock products. Therefore, more and more researchers are turning their attention to developing new, green, and sustainable non-antibiotic feed additives and corresponding alternatives.

[0004] Indole-3-butyric acid (IBA, C12H) 13 NO2 has a wide range of applications: In agriculture, it can induce parthenocarpy in crops, forming seedless fruits, significantly promoting adventitious root formation, rooting of cuttings, and seed germination. Compared with other indole compounds, it is more effective in combating oxidative damage to plants and can also alleviate free radical damage to cell membranes by improving plant stress resistance and reducing reactive oxygen species. In medicine, it can provide new directions for targeted intervention in the treatment of coronary heart disease by regulating tryptophan metabolism balance, and can also induce programmed cell death in colon cancer cells by binding to related receptors and activating downstream pathways, potentially for use in colon cancer treatment. In the animal sector, it can activate the antioxidant defense system of multiple organs in rats and alleviate lipid peroxidation damage to thyroid cell membranes in pigs, showing potential as a natural antioxidant protectant. In aquaculture, the addition of IBA can improve the growth performance of pearl grouper, alleviate intestinal inflammation, and maintain intestinal flora balance. However, its related effects in animal husbandry are rarely reported. The only existing research shows that dietary supplementation with indole-3-acetaldehyde can improve intestinal barrier function by promoting the proliferation of intestinal stem cells. Meanwhile, previous studies of this invention have shown that dietary supplementation with IAA can improve the growth performance of piglets, alleviate diarrhea, enhance antioxidant capacity, and reshape the gut microbiota by regulating gut microbiota.

[0005] However, the specific role of IBA in animal production remains unknown. Summary of the Invention

[0006] In view of the problems existing in the prior art, the present invention aims to explore whether adding IBA to the diet can alleviate diarrhea in weaned piglets, and its effects on growth performance, intestinal health, and intestinal diseases in weaned piglets.

[0007] To achieve the above objectives, the present invention adopts the following technical solution: One objective of this invention is to provide the application of indole-3-butyric acid in the preparation of feed additives, wherein the feed additives have any one or more of the following functions: (1) Alleviate oxidative stress and damage; (2) Improve growth performance; (3) Antidiarrheal; (4) Antioxidant; (5) Improve immune levels; (6) Regulates liver function; (7) Improve gut health; (8) Prevention and treatment of intestinal diseases.

[0008] Furthermore, the improvement in growth performance includes increasing the daily weight gain of animals and / or increasing the apparent digestibility of nutrients.

[0009] Furthermore, the antidiarrheal treatment includes reducing the diarrhea rate in animals, reducing the activity of diamine oxidase in animal serum, and / or reducing the content of D-lactic acid in animal serum.

[0010] Furthermore, the intestinal diseases include colitis.

[0011] Furthermore, the feed additives include pig feed additives.

[0012] The second objective of this invention is to provide a feed additive for improving animal intestinal health and preventing intestinal diseases, which contains indole-3-butyric acid.

[0013] Furthermore, the intestinal diseases include colitis.

[0014] Furthermore, the animals mentioned include pigs.

[0015] The third objective of this invention is to provide a feed for improving animal gut health and preventing intestinal diseases, comprising a basal diet and the aforementioned feed additives.

[0016] Furthermore, the animals mentioned include pigs.

[0017] Compared with the prior art, the present invention has the following beneficial effects: 1. This invention, through in vitro experiments, demonstrates for the first time that IBA has no cytotoxic effects and can alleviate oxidative stress and inflammatory damage to cells within effective concentrations. This lays the groundwork for further research into the biological roles of IBA in animal husbandry and gut health.

[0018] 2. IBA is a natural plant growth regulator. This invention fills the gap in the application of IBA as an indole derivative in livestock production, and provides an experimental basis and theoretical support for IBA to be used as a novel green additive to replace antibiotics and zinc oxide.

[0019] 3. In particular, this invention has also discovered the positive effects of IBA in anti-diarrheal, repairing intestinal barrier damage, relieving intestinal inflammation, and preventing and treating intestinal-related diseases. Attached Figure Description

[0020] Figure 1 This study investigates the effects of different concentrations of IBA treatment for 12 h and 24 h, followed by hydrogen peroxide treatment for 5 h, on the activity of IPEC-J2 cells in porcine intestinal epithelial cells.

[0021] Figure 2 The IBA in this invention targets the antioxidant enzymes in IPEC-J2 cells and AKT The effect of relative mRNA levels.

[0022] Figure 3 This is a statistical analysis of the survival rates of mice in each group during DSS-induced colitis in this invention.

[0023] Figure 4 This invention relates to the effect of IBA on the pathological symptoms of DSS-induced colitis in mice.

[0024] Figure 5 This is a histopathological analysis of colon tissue from different groups in DSS-induced colitis in mice, as described in this invention.

[0025] Figure 6 This invention relates to the effect of IBA on the expression levels of intestinal inflammatory factors in DSS-induced colitis in mice.

[0026] Figure 7 This invention relates to the effect of IBA on the levels of inflammatory factors in mouse serum and colon tissue during DSS-induced colitis.

[0027] Figure 8 This study investigates the effects of DSS-induced colitis in mice on the expression of intestinal tight junction protein factors, mucin factors, and AHR.

[0028] Figure 9 This invention relates to the effects of IBA on the levels of antioxidant enzymes and myeloperoxidase in the colon tissue of different groups of mice.

[0029] Figure 10 This invention relates to the effect of IBA on the expression level of tight junction proteins in DSS-induced colitis. Detailed Implementation

[0030] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the invention. Any modifications or substitutions made to the methods, steps, or conditions of the present invention without departing from the spirit and essence of the invention are within the scope of the invention. The reagents, products, and instruments used in the following examples are all commercially available, and the methods used in the examples, unless otherwise specified, are consistent with conventional methods.

[0031] The technical solution of the present invention will be further described in detail below with reference to the embodiments.

[0032] Example 1: Effects of IBA on H2O2-induced oxidative stress in porcine intestinal epithelial cells IPEC-J2 1. Materials and Methods 1.1 Preparation of complete culture medium DMEM / F12 medium was mixed with premium fetal bovine serum (FBS) at a ratio of 9:1, followed by the addition of 0.1% penicillin-streptomycin solution. The DMEM / F12 complete medium was used to culture porcine jejunal epithelial cells (IPEC-J2).

[0033] 1.2 Preparation of IBA solutions of different concentrations IBA solids were first dissolved to form a stock solution, which was then diluted with DMEM / F12 and RPMI-1640 basal media to prepare IBA solutions with concentrations of 0.25 mM, 0.5 mM, 1 mM, 1.25 mM and 1.50 mM, respectively.

[0034] 1.3 Cell resuscitation, culture and passage Porcine jejunal epithelial cells were cultured overnight using IPEC-J2. Once the intercellular space was below 5% or the cells had reached confluence, they could be passaged. Subsequent experiments could be performed after passages 3-5.

[0035] 1.4 Cell viability assessment To assess the cytotoxicity of IBA, IPEC-J2 cells in logarithmic growth phase were seeded into four 96-well cell culture plates containing the corresponding complete culture medium and cultured at 37°C with 5% CO2 for 24 h, until microscopic observation showed that the cells largely adhered to and covered the bottom of the wells. The culture medium was then discarded, and the cells were treated with a series of prepared IBA solutions of different concentrations for 12 h and 24 h, respectively. After discarding the culture medium, the first 96-well plate corresponding to the 12 h and 24 h values ​​was continued to be cultured in the corresponding basal medium, while the second plate was treated with 1 mM hydrogen peroxide solution for 5 h. A control group was set up in each 96-well plate, using the corresponding sterile culture medium throughout the entire culture process. Finally, cell viability was assessed using the CCK-8 cell proliferation / cytotoxicity assay kit.

[0036] 1.5 Construction of H2O2-induced cellular oxidative stress model and LPS-induced cellular inflammatory damage model To investigate whether IBA could alleviate H2O2-induced damage to IPEC-J2 cells, cells from passages 3-5 were selected for the experiment. Cells from the culture dishes were seeded into six-well plates, ensuring a relatively uniform cell density in each well. After the cells had largely covered the bottom of the wells, they were cultured in IBA solution for 24 h, followed by H2O2 treatment for 5 h. The control group wells contained the corresponding culture medium throughout the experiment.

[0037] 1.6 Collection of Cell Samples After the cell model is successfully constructed, cell samples are collected for subsequent mRNA extraction and assay.

[0038] 1.7 Analysis of the relative expression levels of cytokine mRNA Antioxidant enzymes in IPEC-J2 cells were measured using RT-qPCR. SOD , GSH-Px , CAT and AKT mRNA expression levels.

[0039] 2. Results and Analysis 2.1 Effects of IBA on IPEC-J2 activity in porcine intestinal epithelial cells from Figure 1 As shown in Figure A, after 12 hours of treatment with IBA, IBA showed no toxicity to cells within a certain concentration range (0.25-1 mM), and cell viability tended to increase with increasing IBA concentration. However, high concentrations of IBA (1-1.5 mM) had a certain inhibitory effect on cell proliferation. Figure 1B shows that after 24 h of IBA treatment, IBA significantly improved cell viability in a dose-gradient manner within an appropriate concentration range. This indicates that appropriate concentrations of IBA can promote the growth of IPEC-J2 cells, and the effect becomes more pronounced with increasing culture time. After treating cells with IBA for 12 h and 24 h, respectively, H2O2 was used to intervene in the cells for 5 h. The results showed that the cell viability in the H2O2 group decreased significantly, possibly indicating that H2O2 caused cell damage to some extent. Within the effective concentration range of IBA (0.25-1 mM), IBA can alleviate the toxic effects of H2O2 on cells. Furthermore, the mitigation effect is more pronounced with longer pre-culture time with IBA. After 5 h of hydrogen peroxide treatment, the average relative viability percentage of cells cultured with 0.25-1 mM IBA for 12 h was 96.44%, while the average relative viability percentage of cells cultured with IBA for 24 h was 92.99%. Therefore, it can be concluded that IBA can improve cell viability, promote cell proliferation, and alleviate the toxic damage caused by H2O2 to cells.

[0040] 2.2 Effects of IBA on Indicators Related to Oxidative Stress Injury in IPEC-J2 Cells Induced by Hydrogen Peroxide Figure 2 The results showed that, compared with the control group, the H2O2 group had higher levels of antioxidant enzymes. SOD , GSH-Px , CAT and AKT The mRNA expression level was significantly reduced ( P <0.05). Compared with the H2O2 group, the IBA + H2O2 group SOD , GSH-Px , CAT and AKT The mRNA expression level increased significantly ( P <0.05). Among them GSH-Px The expression level was significantly increased ( P <0.05). This indicates that H2O2 can cause oxidative stress damage to cells, while IBA can alleviate oxidative stress damage caused by hydrogen peroxide by increasing the expression of genes corresponding to antioxidant enzymes.

[0041] Example 2: Effects of indole-3-butyric acid on growth performance, apparent nutrient digestibility, and immune indicators in weaned piglets. 1. Materials and Methods 1.1 Experimental Design and Experimental Animals This experiment selected 240 healthy weaned piglets of Duroc × Landrace × Large White crossbred, aged 28 days. The experiment followed a randomized complete block design, randomly dividing the weaned piglets into 5 treatment groups based on weight, sex, and equal numbers of males and females. Each treatment group had 6 replicates (pens), with 8 piglets per replicate. The entire experiment lasted 28 days. The positive control group was fed a corn-soybean meal basal diet containing zinc oxide for the first 14 days and without zinc oxide for the following 14 days. The blank control group was fed a corn-soybean meal basal diet without zinc oxide. The experimental groups had 200, 400, and 800 mg / kg of IBA added to their basal diets, respectively.

[0042] 1.2 Experimental Diets and Feeding Management Before the formal start of the experiment, weaned piglets were transferred to the nursery and fed creep feed for 1-3 days as a transition. During the experiment, the health and mental state of piglets in each pen were carefully observed and recorded daily. During the experiment, the excretion of all piglets in each pen was observed twice daily, at 9:00 AM and 3:00 PM, and the corresponding diarrhea rate was statistically analyzed. The corn, bran, and soybean meal in the experimental diet were provided by the animal experimental base, while other feed ingredients were purchased from Beijing Tonglixingke Agricultural Technology Co., Ltd. Various amino acid raw materials, limestone powder, salt, and other additives were weighed, packaged, and labeled according to their processing requirements at the Pinggu feed farm of Beijing Tonglixingke Agricultural Technology Co., Ltd., and then transported to the Fengning base. Finally, the feed powder was prepared by a series of processing steps, including formulation, crushing, and mixing, in the feed pilot plant of the animal experimental base. The basal diet was formulated according to the national standard GB / T 39235-2020 Nutritional Requirements for Pigs (8-25 kg piglets) to meet the nutritional needs of the piglets. The basal diet did not contain antibiotics; the specific formula and ingredient composition are shown in Table 1.

[0043] 1.3 Sample Collection During the last three days of the experiment, approximately 300 g of fresh feces from each pen of piglets was collected in the morning and afternoon, mixed thoroughly, and aseptically stored in a -20°C cold storage. After the experiment, the feces were dried, pulverized, and used to determine the apparent intestinal digestibility of nutrients.

[0044] On days 14 and 28 of the experiment, one piglet was randomly selected from each pen. After restraint, 6-7 mL of blood was collected from the anterior vena cava and placed in a vacuum blood collection tube without anticoagulant. After standing at room temperature for 40 min, the blood was centrifuged at 3000 rpm / min and 4℃ for 20 min. The serum was collected and aliquoted into 2 mL centrifuge tubes and stored at -20℃ for subsequent determination of various indicators.

[0045] 1.4 Detection Indicators and Measurement Methods (1) Growth performance On days 0, 14, and 28 of the experiment, piglets were weighed in pens to calculate the average daily gain (ADG) at different time points (0-14 days, 15-28 days, 0-28 days). Piglets were fasted for 12 hours before weighing. The remaining feed in the feed troughs of each pen was observed regularly, and the feed consumption for each pen was recorded based on the number of feedings to calculate the average daily feed intake (DFI). The feed conversion ratio (F:G) was finally calculated based on the average daily feed intake and average daily gain. The calculation formulas are as follows: Average daily gain (ADG) = (Final weight of piglets - Initial weight of piglets) / Number of days in the experiment; Average daily feed intake (ADFI) = Feed consumption / (Number of piglets per group × Number of days in the experiment); Feed conversion ratio (F / G) = Average daily feed intake / Average daily gain.

[0046] (2) Stool score and diarrhea rate During the experiment, the piglets' feed intake, water consumption, and feces were observed and recorded twice daily, at 9:00 AM and 3:00 PM. Feces from each pen were randomly scored using a 3-point scoring system: 0 points for normal feces (long, cylindrical, soft, elastic, and smooth); 1 point for soft feces; 2 points for feces becoming thin and pasty and unable to maintain a fixed shape; and 3 points for watery, shapeless, and highly fluid feces. If a piglet scored 2 or 3 points for two or more consecutive days, it was considered to have diarrhea. The diarrhea rate was calculated as follows: Piglet diarrhea rate (%) = Number of piglets with diarrhea during the experiment / (Number of piglets per group × Number of days in the experiment) × 100.

[0047] (3) Determination of apparent intestinal digestibility of nutrients The collected feed ingredients and fecal samples were thoroughly ground and mixed evenly through a 1 mm sieve for the determination of apparent nutrient digestibility. Various nutrients in the test diet and feces, including dry matter (DM), crude fat (EE), crude ash (Ash), crude protein (CP), crude fiber (CF), neutral detergent fiber (NDF), acid detergent fiber (ADF), and total energy (GE), were determined using the endogenous indicator method of hydrochloric acid-insoluble ash. Hydrochloric acid-insoluble ash was determined according to GB / T 23742-2009. Crude fat (EE), crude ash (Ash), crude protein (CP), and dry matter (DM) were determined according to GB / T 6433-2025, GB / T 6438-2025, GB / T 6432-2018, and GB / T 6435-2014. Calcium and phosphorus were determined according to the methods in GB / T 6436-2018 and GB / T 6437-2018, respectively. Crude fiber (CF), neutral detergent fiber (NDF), and acid detergent fiber (ADF) were determined according to the methods in GB / T 6434-2022, GB / T 20806-2022, and NY / T 1459-2022, respectively. The total energy (GE) of all samples was determined using an oxygen bomb calorimeter. The apparent digestibility of various nutrients was calculated using an indicator method, with the following formula: Apparent digestibility of a nutrient (%) = 100% - (AIA content in the diet / AIA content in the fecal sample × nutrient content in the fecal sample / nutrient content in the diet) × 100%.

[0048] (4) Measurement of serum markers Serum levels of inflammatory factors TNF-α, IL-1β, IL-6, and IL-10, as well as immunoglobulin levels IgM, IgA, and IgG, were measured using enzyme-linked immunosorbent assay (ELISA). Serum glutathione peroxidase (GSH-Px) activity and D-lactic acid content were measured colorimetrically. Serum superoxide dismutase (SOD) activity was measured using the xanthine oxidase method. Serum total antioxidant capacity (T-AOC) and diamine oxidase (DAO) activity were measured using the ABTS microplate method. For specific procedures regarding serum aspartate aminotransferase (AST), alkaline phosphatase (ALP), and alanine aminotransferase (ALT), please refer to the respective kit instructions.

[0049] Table 1. Composition and nutrient levels of the basal diet (%, feeding basal level) 1

[0050] Note: The basal diet formulation for the positive control group from day 0 to 28 is shown in the table above. The basal diet formulations for the blank control group and the IBA treatment group from day 0 to 28 are the same as those for the positive control group from day 14 to 28. Different concentrations of IBA treatment groups had 200, 400, and 800 mg / kg of IBA-Na (purity > 98%) additionally added to their basal diet formulations. IBA-Na was purchased from Wuhan Jiangxin Biotechnology Co., Ltd. 2 The soybean oil was purchased from Beijing Fuchenxinsheng Trading Co., Ltd. 3 Sucralose was purchased from Anhui Jinhe Industrial Co., Ltd. 4 The premix provides the following vitamins and minerals per kilogram of diet: Vitamin A, 12,000 IU; Vitamin B1, 2.5 mg; Vitamin B2, 4 mg; Vitamin B6, 3 mg; Vitamin B12, 0.02 mg; Vitamin D3, 2,500 IU; Vitamin E, 30 IU; Vitamin K3, 2.5 mg; Pantothenic acid, 12.5 mg; Biotin, 0.14 mg; Folic acid, 0.7 mg; Nicotinamide, 40 mg; Zinc, 0.09 g; Iron, 0.3 g; Iodine, 0.5 mg; Copper, 0.09 g; Selenium, 0.33 mg; Manganese, 0.05 g.

[0051] 2. Results and Analysis 2.1 Effects of IBA on growth performance and diarrhea rate in weaned piglets The effects of dietary IBA supplementation on the growth performance and diarrhea rate of weaned piglets are shown in Table 2. The results indicated that, compared to the control group, the 200 mg / kg and 400 mg / kg IBA treatment groups increased the average daily weight gain of piglets from day 0 to 14 of the experiment. However, different doses of IBA did not significantly differ in average daily gain (ADG), average daily feed intake (ADFI), feed conversion ratio (F:G), or body weight at different time points. Meanwhile, from day 15 to 28 of the experiment, there was no significant difference in the diarrhea rate among the treatment groups, but the 400-800 mg / kg IBA treatment groups showed a significant trend towards reducing the diarrhea rate (<0.05). P <0.01). During days 0-14 and throughout the entire experimental phase 0-28, compared to the control group, the addition of 200-800 mg / kg IBA to the diet significantly reduced the diarrhea rate in piglets. P <0.05).

[0052] The above results indicate that adding different concentrations of IBA to the diet significantly reduced the diarrhea rate in piglets in a dose-dependent manner during the early stage of the experiment (days 0-14) and throughout the entire experimental period (days 0-28). P<0.05), and IBA can linearly reduce the diarrhea rate in weaned piglets (linear, P = 0.004, P = 0.009). Overall, compared to other doses of IBA, adding 800 mg / kg of IBA to the diet was the most effective in reducing the diarrhea rate in weaned piglets.

[0053] Table 2. Effects of IBA on growth performance and diarrhea rate in weaned piglets. 1

[0054] Note: Different letters in the same line a-b This indicates that there are significant differences between the different treatment groups. P <0.05). Positive control group CON1: Zinc oxide 2000 mg / kg was added to the basal diet for the first 14 days, and no zinc oxide was added for the next 14 days; Blank control group CON2: No zinc oxide was added to the basal diet; IBA treatment group: IBA was added to the basal diet without zinc oxide by 200, 400, and 800 mg / kg, respectively. 2 n = 6. 3 DI: Diarrhea incidence = (Total number of piglets with diarrhea during the trial period) / (Total number of pigs × Total number of trial days) × 100.

[0055] 2.2 Effects of IBA supplementation on the apparent digestibility of various nutrients in diets The effects of zinc oxide supplementation and different doses of IBA on the apparent digestibility of nutrients in weaned piglets are shown in Table 3. There were no significant differences in the apparent digestibility of crude fiber among the treatment groups. Compared with the control group, the 200 mg / kg IBA treatment group significantly increased the ATTD (attempted digestibility) of calcium (Ca), phosphorus (P), crude ash (Ash), and neutral detergent fiber (NDF). P <0.05), there were no significant differences in ATTD for other nutrients; the 400 mg / kg IBA treatment group increased the ATTD of most nutrients, with significant differences in ATTD for calcium (Ca), phosphorus (P), dry matter (DM), total energy (GE), crude ash (Ash), neutral detergent fiber (NDF), and acid detergent fiber (ADF). P <0.05), while the ATTD of crude protein (CP), crude fiber (CF) and crude fat (EE) were increased respectively, but there was no significant difference; the 800 mg / kg IBA treatment group showed an increasing trend in the ATTD of calcium (Ca), but there was no significant difference in the ATTD of other nutrients.

[0056] The above results indicate that adding 200-400 mg / kg of IBA to the diet can significantly improve the apparent digestibility of nutrients, with the 400 mg / kg IBA treatment group showing better results. However, the 800 mg / kg IBA treatment group did not show significant differences in the ATTD of most nutrients, and even reduced the ATTD of some nutrients, suggesting that adding high doses of IBA to the diet may have a negative effect on nutrient digestibility.

[0057] Table 3 Effect of IBA on apparent digestibility of nutrients in weaned piglets (%, dry basis)

[0058] Note: Different letters "ab" in the same row indicate significant differences between different treatment groups (P<0.05). Positive control group CON1: Zinc oxide 2000 mg / kg was added to the basal diet for the first 14 days, and no zinc oxide was added for the next 14 days; Blank control group CON2: No zinc oxide was added to the basal diet; IBA treatment groups: IBA was added to the basal diet without zinc oxide by 200, 400, and 800 mg / kg, respectively. n = 6.

[0059] 2.3 Effects of IBA on serum immunoglobulins in weaned piglets The effects of dietary supplementation with different doses of IBA on the levels of immunoglobulins IgA, IgM, and IgG in the serum of weaned piglets are shown in Table 4. As can be seen from the table, compared with the blank control group, the 200-800 mg / kg IBA treatment groups increased the levels of IgA and IgG in the serum of piglets on day 28 and the level of IgM in the serum of piglets on day 14, but there were no significant differences between the treatment groups. Compared with the blank control group, the 800 mg / kg IBA treatment groups significantly increased the levels of IgA and IgG in the serum of piglets on day 14. P <0.05), the IBA treatment groups of 200-400 mg / kg increased serum IgA and IgG levels on day 14 of the experiment, but there were no significant differences between the treatment groups. Meanwhile, IgA and IgG levels increased significantly in a dose-dependent manner with IBA (linear). P <0.01). Compared with the blank control group, the IBA treatment groups of 200-800 mg / kg significantly increased the level of IgM in the serum of piglets on day 28 of the experiment ( P <0.05), and IBA can linearly increase the level of IgM in the serum of piglets (linear, P <0.01).

[0060] The results indicate that, throughout the experiment, adding different doses of IBA to the diet increased the secretion levels of immunoglobulins IgA, IgM, and IgG in weaned piglets to varying degrees. Overall, 800 mg / kg of IBA showed the best effect.

[0061] 2.4 Effects of IBA on the antioxidant capacity of weaned piglet serum The effects of dietary supplementation with different doses of IBA on the activities of antioxidant enzymes superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and antioxidant capacity (T-AOC) in the serum of weaned piglets are shown in Table 4. The results indicate that, compared with the blank control group, the 200-800 mg / kg IBA treatment group increased the activity of antioxidant enzyme GSH-Px in the serum of piglets on day 28 of the experiment, but there was no significant difference among the treatment groups; while the 200-800 mg / kg IBA treatment group significantly increased the activity of antioxidant enzyme GSH-Px in the serum of piglets on day 14 of the experiment. P <0.05. Compared with the blank control group, the IBA treatment groups of 200 mg / kg and 800 mg / kg significantly increased the total antioxidant capacity of piglet serum on days 14 and 28 of the experiment, respectively. P <0.05), there was no significant difference in the 400 mg / kg IBA treatment group, but the data indicated that IBA could linearly increase the total antioxidant capacity in serum (linear, P <0.05%. Compared with the blank control group, the positive control group significantly increased the activity of SOD in piglet serum on days 14 and 28 of the experiment. P <0.05), there was no significant difference in serum SOD activity among the different IBA treatment groups on day 14 of the experiment, but the serum SOD activity in the 800 mg / kg IBA treatment group showed an increasing trend (<0.05). P <0.10).

[0062] The above results indicate that dietary IBA supplementation can improve the total antioxidant capacity and antioxidant enzyme activity of piglet serum to some extent. Overall, the IBA treatment group with 400-800 mg / kg showed relatively better results.

[0063] 2.5 Effects of IBA on serum biochemical parameters of weaned piglets The effects of dietary supplementation with different doses of IBA on serum biochemical parameters aspartate aminotransferase (AST), alkaline phosphatase (ALP), and alanine aminotransferase (ALT) in weaned piglets are shown in Table 4. The results showed that, compared with the control group, the 200-800 mg / kg IBA treatment groups reduced AST activity on day 14 and day 28, respectively, but there were no significant differences among the treatment groups. Compared with the control group, the 400 mg / kg IBA treatment group significantly reduced serum ALT activity on day 14. P <0.05), the 800 mg / kg IBA treatment group showed a more significant decreasing trend in serum ALT activity on days 14 and 28 of the experiment (<0.05). P <0.10), the 200 mg / kg IBA treatment group reduced ALT activity at both time points, but the difference was not statistically significant. Compared with the blank control group, the positive control group significantly increased serum ASP activity on day 14 of the experiment ( P (<0.05) IBA treatments at 400 mg / kg and 800 mg / kg reduced serum ALP activity, as did the 200 mg / kg IBA treatment group, but none showed significant differences. However, on day 28 of the experiment, all IBA treatments increased serum ALP activity, but there were no significant differences between the treatment groups.

[0064] The above results indicate that on day 14 of the experiment, different doses of IBA could reduce the activity of liver function-related enzymes AST, ALP, and ALT in piglets to some extent, while on day 28 of the experiment, it increased the activity of ALP.

[0065] 2.6 Effects of IBA on intestinal permeability in weaned piglets The effects of dietary supplementation with different doses of IBA on serum diamine oxidase (DAO) and D-lactic acid (D-LA) levels in weaned piglets are shown in Table 4. The results indicated that, compared to the control group, different doses of IBA reduced serum DAO activity on day 28 of the experiment, but there were no significant differences between the treatment groups. The 200-800 mg / kg IBA treatment group significantly reduced serum DAO activity on day 14 of the experiment. P <0.05), and the decrease in DAO exhibits a linear dose-response effect. (Linear, P = 0.007). Compared with the blank control group, on day 14 of the experiment, the 800 mg / kg IBA treatment group significantly reduced serum D-LA levels ( P <0.05), with the 400 mg / kg IBA treatment group showing a decreasing trend in serum D-LA levels (<0.05).P <0.10), the 200 mg / kg IBA treatment group reduced serum D-LA levels, but there was no significant difference between the two groups. Compared with the blank control group, on day 28 of the experiment, the 400 mg / kg IBA treatment group significantly reduced serum D-LA levels ( P <0.05), among which the serum D-LA content in the 800 mg / kg IBA treatment group showed a decreasing trend (0.05 < P <0.10), the 200 mg / kg IBA treatment group reduced the serum D-LA content, but there was no significant difference between the two groups.

[0066] The above results indicate that dietary supplementation with IBA can, to some extent, reduce DAO activity and D-lactic acid levels in piglet serum. Overall, the 400-800 mg / kg IBA treatment group showed relatively better results.

[0067] 2.7 Effects of IBA on serum inflammatory factors in weaned piglets The effects of dietary supplementation with different doses of IBA on serum levels of inflammatory factors TNF-α, IL-1β, IL-6, and IL-10 in weaned piglets are shown in Table 4. The results show that, compared with the control group, the 200-800 mg / kg IBA treatment groups reduced serum TNF-α and IL-6 levels on day 14 and serum IL-1β levels on day 28, but there were no significant differences among the treatment groups. However, on day 28, the 400 mg / kg IBA treatment group significantly reduced serum TNF-α levels (…). P <0.05), serum TNF-α levels in the 200 and 400 mg / kg IBA treatment groups showed a more significant decreasing trend (<0.05). P <0.10, but there was no significant difference; on day 28 of the experiment, the 400-800 mg / kg IBA treatment group significantly reduced serum IL-6 levels ( P <0.05), the serum IL-6 level in the 200 mg / kg IBA treatment group showed a decreasing trend (0.05 < P <0.10), but there was no statistically significant difference. Compared with the blank control group, on day 14 of the experiment, the 800 mg / kg IBA treatment group significantly increased the serum IL-10 level ( P <0.05), serum IL-10 levels tended to increase in the 400 mg / kg IBA treatment group (<0.05). P<0.10) The 200 mg / kg IBA treatment group increased serum IL-10 levels, but the differences were not statistically significant. The data also show that IBA can linearly increase IL-10 levels (linear, P =0.002); on day 28 of the experiment, the IBA treatment groups of 200-800 mg / kg all significantly increased serum IL-10 levels ( P <0.05), and it increased significantly with increasing IBA addition (linear). P <0.001).

[0068] The above results indicate that dietary supplementation with IBA can, to some extent, reduce the levels of pro-inflammatory factors TNF-α, IL-1β, and IL-6 in piglet serum, while increasing the level of the anti-inflammatory factor IL-10, exhibiting a dose-dependent effect. Overall, the IBA treatment group at 400-800 mg / kg showed relatively better results.

[0069] Table 4. Effects of IBA on antioxidant capacity, immune capacity, inflammatory factor levels, intestinal permeability indicators, and serum biochemical indicators in weaned piglets.

[0070] Note: Different letters "ab" in the same line indicate significant differences between different treatment groups. P <0.05). n = 6.

[0071] Example 3: Investigation into the mechanism of indole-3-butyric acid in alleviating DSS-induced colitis in mice. 1. Materials and Methods 1.1 Preparation of IBA solutions of different concentrations administered to mice by gavage Mice were acclimatized for 5 days before gavage. At the end of the 5-day period, the mouse weights were as follows: control group 18.20 ± 0.45 g, DSS group 19.18 ± 0.30 g, low-concentration IBA group 19.06 ± 0.18 g, and high-concentration IBA group 18.22 ± 0.52 g. Each mouse was gavaged with 200 μL of PBS. Based on an average mouse weight of 18 g, the final gavage concentrations of IBA-K were calculated to be 20 mg / kg and 40 mg / kg. High-purity (98%) potassium indole-3-butyrate (IBA-K) was purchased from Shanghai Yuanye Biotechnology Co., Ltd.

[0072] 1.2 Experimental Animals and Their Husbandry Management This experiment used 50 female SPF-grade C57BL / 6J mice aged 6-8 weeks, purchased from Beijing Spaford Biotechnology Co., Ltd. The mice were housed in an SPF-grade barrier environment at the Animal Experiment Platform of the College of Veterinary Medicine, West Campus, China Agricultural University (room temperature: 22-26℃; daily temperature difference: ≤ 4℃; relative humidity: 50%-70%; air exchange rate: 15-20 times / h).

[0073] 1.3 Experimental Design Fifty mice were randomly divided into four groups according to their body weight: a control group, a DSS model group, a low-concentration (20 mg / kg) IBA treatment group, and a high-concentration (40 mg / kg) IBA treatment group, with 10 mice in each group and 5 mice per cage. Mice underwent a 5-day acclimatization period before the experiment, and the formal experiment lasted 16 days. The control and DSS groups were administered 200 μL of sterile PBS buffer by gavage daily, while the IBA treatment groups were administered 200 μL of 20 mg / kg and 40 mg / kg IBA-K solutions by gavage daily. For nine days after the experiment, in addition to the corresponding solutions administered by gavage, the DSS and IBA treatment groups were given water containing 2.5% DSS daily, with the drinking water changed every three days. Mice had free access to food and water during the experiment.

[0074] 1.4 Sample Collection Mice were weighed 24 hours after administration on the last day of the experiment, marking the end of the experiment. All mice were euthanized by cervical dislocation after blood was collected from their eyes, followed by dissection and sampling. The collected serum was allowed to stand at room temperature for 5 hours, then centrifuged at 3000 rpm for 15-20 minutes at 4°C and stored at -80°C for subsequent determination of relevant indicators. The colonic tissue was completely removed, its length measured, and photographed. The colonic tissue was longitudinally dissected with scissors, carefully washed in a culture dish containing sterile PBS buffer to remove residual food residue and blood, and cut into multiple segments of 1.5-2 cm in length. One segment was collected and fixed in 4% paraformaldehyde for subsequent histopathological analysis of the colonic tissue; another segment was placed in a 1.5 mL sterile centrifuge tube, flash-frozen in liquid nitrogen, and then transferred to a -80°C freezer for long-term storage for subsequent determination of cytokine and related protein expression levels.

[0075] 1.5 Detection Indicators and Measurement Methods (1) Mouse survival rate, body weight and mental status During the experiment, specifically the first 7 days after IBA intervention and the last 9 days after DSS treatment, the mice were weighed at the same time each day to observe their physical condition, mental state, and survival rate. Figure 3 ).

[0076] (2) Disease Activity Index (DAI) Nine days after the start of the experiment involving the addition of DSS to drinking water, the mice's condition was observed daily, including recording their weight and Disease Activity Index (DAI). DAI comprised three aspects: percentage weight loss, degree of fecal occult blood, and fecal consistency. A cumulative score was calculated based on these three indicators: DAI = Percentage Weight Loss Score + Fecal Occult Blood Score + Fecal Characteristics Score. The scoring details for the Disease Activity Index followed Qin's method. Fecal occult blood levels were detected using a fecal occult blood test kit; detailed procedures are provided in the instruction manual. Scoring details and indicators are shown in Table 6.

[0077] (3) Morphological and pathological analysis of mouse colon tissue A 1.5 cm segment of colonic tissue was collected and fixed in 4% paraformaldehyde. After embedding in paraffin, colonic tissue sections were prepared. The sections were stained with hematoxylin and eosin (HE) and observed under a light microscope for histopathological morphology and analysis. The colonic histopathological scoring system followed Mosli's scoring system, which assessed nine aspects: degree of erosion / ulceration, severity of inflammation, fibrosis, crypt damage, lesion depth, extent of erosion / ulceration, inflammatory cell infiltration, goblet cell reduction, and tissue damage. Each indicator was scored from 0 to 4 points based on its severity. The final histopathological score was the sum of the scores for each indicator; detailed scoring rules are shown in Table 5.

[0078] (4) Determination of relative expression levels of cytokine mRNA in mouse colon tissue By observing colon tissue sections and histopathological analysis results, total mRNA was extracted from pre-collected colon tissue fragments, and tight junction proteins in the colon tissue were detected using RT-qPCR. ZO-1 , Claudin and Occludin ), inflammatory cytokines ( TNF-α , IL-1β , IL-6 and IL-10 ) and mucin factor ( Mucin-1 , Mucin-2 The relative expression levels of IBA mRNA were measured. To investigate whether IBA is a specific ligand for the aryl hydrocarbon receptor (AHR), colon tissue was also examined. AHR Gene expression levels.

[0079] (5) Determination of cytokines in mouse serum After removing the serum from the -80°C freezer and allowing it to thaw completely, centrifuge it again and measure cytokines using enzyme-linked immunosorbent assay (ELISA). TNF-α , IL-1β , IL-6 and IL-10 .

[0080] (6) Determination of antioxidant enzyme, MPO and MDA content in mouse colon tissue Colonic tissue was removed from a -80°C freezer and placed on ice. A certain amount of tissue was weighed and washed with sterile PBS buffer to remove blood, and the weight of each sample was recorded. The tissue was cut into small pieces and placed in a tissue homogenizer, with appropriate grinding beads and 1 mL of 1× PBS solution added. After homogenization, the homogenate was placed in a -20°C freezer overnight. After repeated freeze-thaw cycles to disrupt cell membranes, the tissue homogenate was centrifuged at 2-8°C at 5000×g for 5 minutes, and the supernatant was collected for subsequent experiments. The activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and myeloperoxidase (MPO) in colonic tissue were detected using an ELISA kit; the content of malondialdehyde (MDA) in colonic tissue was detected using the thiobarbituric acid (TBA) method.

[0081] (7) Measurement of cytokines in mouse colon tissue The levels of inflammatory factors TNF-α, IL-1β, IL-6 and IL-10 in colon tissue were detected using an ELISA kit.

[0082] (8) Determination of tight junction protein expression levels in mouse colon tissue The expression levels of the tight junction proteins ZO-1, Claudin, and Occludin in colon tissue were detected using Western blotting.

[0083] Table 5. Scoring criteria for colonic histopathology in mice with DSS-induced colitis.

[0084] Table 6 Disease Activity Index (DAI) Scoring Table

[0085] 2. Results and Analysis 2.1 IBA alleviated DSS-induced colitis in mice Throughout the experiment, mice in both the control and DSS model groups were administered sterile PBS buffer via gavage daily. Before and after DSS challenge, mice in the IBA treatment groups were administered equal amounts of 20 mg / kg and 40 mg / kg IBA via gavage daily. Daily observation of the mice revealed that the control group mice were generally more active, ate normally, had glossy fur, and interacted frequently with each other. The health and mental state of mice in both the low- and high-concentration IBA treatment groups were affected to varying degrees, with reduced activity and decreased motor function; the effects were more pronounced in the low-concentration IBA treatment group. Mice in the DSS model group exhibited depression, sluggishness, dull eyes, and a dull, disheveled coat. They also displayed abnormal postures such as arched backs and ruffled fur, and further observation revealed uncoordinated movements such as a staggering gait. These symptoms worsened with prolonged DSS challenge. At the end of the experiment, some mice in the DSS model group even became immobile and lost all motor function.

[0086] As shown in Table 7, the body weight of mice decreased continuously during DSS administration. The DSS group mice experienced a more significant weight loss starting on day 5, with a decrease of approximately 15% by the last day. IBA treatment alleviated the DSS-induced weight loss, with 40 mg / kg IBA treatment showing a more significant improvement in body weight. P <0.05). Simultaneously, the fecal condition of each group of mice was observed during DSS modeling. The feces of the DSS group mice were relatively loose and unformed, with some mice's feces adhering to the cage walls. After day 6, blood in the feces and anal bleeding were visible to the naked eye. The bloody stool and unformed feces of the IBA group mice were alleviated. From Figure 4 As shown in (B), compared with the DSS model group, the IBA treatment group can alleviate the increase in DAI score caused by DSS to a certain extent. The 40 mg / kg IBA group significantly reduced the DAI score. P <0.05). Finally, the DSS model group mice showed shortened colons and enlarged spleens. Compared with the DSS model group, the 40 mg / kg IBA group significantly inhibited colon shortening and reduced spleen weight to body weight ratio. P <0.05).

[0087] Table 7 Effects of IBA on body weight changes in DSS-induced colitis in mice

[0088] Note: Different letters in the same line a-b This indicates that there are significant differences between the different treatment groups. P<0.05). Control group CON: Orally administered sterile PBS buffer daily during the experiment; DSS group: Orally administered an equal volume of sterile PBS buffer daily before DSS induction, and drinking water containing 2.5% DSS during DSS induction; IAA_20 group and IAA_40 group: Orally administered 20 mg / kg and 40 mg / kg IBA-K daily before and after DSS induction. 2 n = 9.

[0089] 2.2 Effects of IBA on DSS-induced colonic tissue damage in mice HE-stained sections were examined under a light microscope for colonic pathological morphology and histopathological analysis. Results showed that the colonic tissue of control mice showed no pathological damage, with abundant and densely arranged lamina propria crypts and goblet cells. However, the colonic tissue of DSS-induced mice exhibited significant inflammatory damage. Figure 5 In the DSS model group of mice, extensive ulceration of the colon tissue was observed, with loss of mucosal epithelium and crypt structures, severe reduction of goblet cells, and extensive connective tissue hyperplasia accompanied by infiltration of numerous lymphocytes and granulocytes. The submucosa showed minimal connective tissue hyperplasia, vasodilation, and significant lymphocyte infiltration. The lamina propria showed minimal crypt dilation, flattened crypt epithelial cells, and eosinophilic material within the glandular lumen. The muscular layer exhibited uneven thickness. The intestinal lumen contained a small amount of necrotic cell debris, sloughed epithelial cells, erythrocytes, and eosinophilic material. IBA treatment alleviated the pathological damage to the colon tissue caused by DSS, with the 40 mg / kg IBA treatment group showing the best improvement, significantly reducing the pathological score of the mouse colon tissue. P <0.05), which alleviated DSS-induced colonic tissue damage in various ways, specifically manifested as rare colonic ulcers less than 3 mm, disappearance of basal 1 / 3 crypt structure, mild reduction of goblet cells, a small amount of connective tissue hyperplasia, accompanied by a small amount of lymphocyte infiltration; inflammatory cells infiltrated into the submucosa; a small amount of lymphocytes focally aggregated in the lamina propria; and uneven thickness of the muscular layer.

[0090] 2.3 Effects of IBA on DSS-induced inflammatory response in colonic tissue Detection of inflammatory cytokines in mouse colon tissue using RT-qPCR TNF-α , IL-1β , IL-6 and IL-10 The relative expression levels of mRNA, the results are as follows Figure 6 Compared with the control group, the DSS model group mice had higher levels of anti-inflammatory factors in their colon tissue. IL-10 The mRNA expression level of pro-inflammatory factors was significantly reduced. TNF-α , IL-1β and IL-6 The expression level of decreased significantly ( P<0.05). Compared with the DSS model group, the 20 mg / kg IBA treatment group significantly reduced pro-inflammatory factors. TNF-α and IL-1β mRNA expression level ( P <0.05), the 40 mg / kg IBA treatment group significantly improved IL-10 The mRNA expression level of [the substance] was significantly reduced, while pro-inflammatory factors were also significantly decreased. TNF-α , IL-1β and IL-6 mRNA expression level ( P <0.05).

[0091] The levels of inflammatory factors in colon tissue were detected using an ELISA kit, and the overall trend and results were largely consistent with those of RT-qPCR. Figure 7 As can be seen from (EH), compared with the control group, the levels of TNF-α, IL-1β, and IL-6 in the colon tissue of mice in the DSS model group were significantly increased, while the level of IL-10 was significantly decreased. P <0.05). Compared with the DSS model group, treatment of mice with different concentrations of IBA significantly reduced the secretion levels of TNF-α, IL-1β, and IL-6 in colonic tissue to varying degrees, while significantly increasing [the levels of other substances]. IL-10 secretion ( P <0.05).

[0092] This experiment established a DSS-induced mouse colitis model. The results showed that after DSS administration, the levels of pro-inflammatory factors in the mouse intestinal tissue increased significantly, indicating that DSS can cause a severe reaction in the mouse intestinal tissue. IBA treatment in mice can alleviate the DSS-induced colitis to some extent, with 40 mg / kg of IBA showing better efficacy and a more significant inhibitory effect on the inflammatory response.

[0093] 2.4 Effects of IBA on DSS-induced intestinal mucosal barrier damage in mice Detection of tight junction proteins in mouse colon tissue using RT-qPCR ( ZO-1 , Claudin , Occludin ) and mucin factor ( Mucin-1 , Mucin-2 The relative expression levels of mRNA of [a specific group] are shown in the figure. From [the data]... Figure 8 As can be seen, compared with the control group, the expression levels of tight junction protein and mucin factor mRNA in the DSS model group mice were significantly reduced. PThe value <0.05 indicates that DSS-induced colitis in mice caused intestinal damage and disruption of the intestinal mucosal barrier. Compared with the DSS model group, the IBA treatment group could restore the gene expression of related factors. Specifically, the 20 mg / kg IBA treatment group showed improved gene expression of related factors. Occludin and ZO-1 There was an increasing trend in mRNA expression levels, which significantly improved... Mucin-1 Gene expression ( P <0.05); while the 40 mg / kg IBA treatment group showed... Mucin-1 The expression level showed an increasing trend, while significantly improving... Occludin and ZO-1 Gene expression ( P <0.05).

[0094] The results above show that IBA can increase the expression of intestinal tight junction proteins, alleviate intestinal damage caused by DSS, and restore the integrity of the intestinal mucosa to a certain extent.

[0095] 2.5 Effects of IBA on antioxidant enzymes, MPO activity, and MDA content in DSS-induced mouse colon tissue The activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), myeloperoxidase (MPO), and the content of malondialdehyde (MDA) in colon tissue were detected using an ELISA kit and TBA method. The results are as follows: Figure 9 Inflammatory response and oxidative stress damage are inextricably linked. As shown in the figure, compared with the control group, the SOD level in the DSS model group showed a decreasing trend in the colon, and the GSH-Px level was significantly reduced. P <0.05); Myeloperoxidase (MPO) activity indicates the degree of neutrophil infiltration. Under normal circumstances, the content of myeloperoxidase in monocytes is low. Compared with the control group, the MPO activity in the DSS model group was significantly increased ( P <0.05); Lipid oxidation occurs in animal or plant cells during oxidative stress. Some fatty acids, after oxidation, gradually decompose into a series of complex compounds, including MDA. Compared to the control group, the MDA content in the colon of the DSS model group was significantly increased ( P <0.05). However, compared with the DSS model group, both the 20 mg / kg and 40 mg / kg IBA treatment groups significantly reduced the content of MDA in colonic tissue and significantly reduced the activity of MPO ( P <0.05; the 20 mg / kg IBA treatment group significantly restored the level of SOD in the intestine ( P <0.05), in GSH-PxThe level showed an increasing trend; the 40 mg / kg IBA treatment group significantly restored the level of GSH-Px in the intestine ( P <0.05), indicating an increasing trend in SOD levels.

[0096] The results above show that DSS-induced colitis in mice not only causes intestinal inflammation but also involves oxidative stress damage. IBA can alleviate the oxidative stress damage caused by DSS, increase the level of antioxidant enzymes in the intestine, reduce MPO activity, and decrease the content of MDA in the intestine.

[0097] 2.6 Effect of IBA on the levels of inflammatory factors in the serum of mice induced by DSS The levels of TNF-α, IL-1β, IL-6, and IL-10 in mouse serum were determined using an ELISA kit. The results are as follows: Figure 7 (AD), as shown in the figure, compared with the control group, the serum levels of inflammatory factors TNF-α, IL-1β and IL-6 in mice in the DSS model group were significantly increased. P <0.05), while anti-inflammatory factors IL-10 There was an increasing trend. Compared with the DSS model group, the 20 mg / kg IBA treatment group significantly increased the level of the anti-inflammatory factor IL-10 in mouse serum, while significantly decreasing the level of IL-10 in serum. TNF-α , IL-1β , IL-6 content ( P <0.05; the 40 mg / kg IBA treatment group significantly increased serum IL-10 levels, while significantly decreasing the levels of TNF-α, IL-1β, and IL-6. P <0.05).

[0098] The above results are basically consistent with the expression trends of inflammatory factors in mouse colon tissue, further indicating that IBA can alleviate DSS-induced inflammatory response, reduce the levels of pro-inflammatory factors TNF-α, IL-1β and IL-6, and increase the level of anti-inflammatory factor IL-10.

[0099] 2.7 Effect of IBA on the expression level of tight junction protein in mouse colon tissue To further clarify the role of IBA in DSS-induced intestinal barrier function in mice, Western blotting was used to determine the expression levels of tight junction proteins ZO-1, Claudin, and Occludin in the mouse colon. Figure 10 It can be seen that, compared with the control group, the expression levels of ZO-1, Claudin, and Occludin in the colon of mice in the DSS model group were significantly decreased. P<0.05. Compared with the DSS model group, the 20 mg / kg and 40 mg / kg IBA treatment groups restored the expression levels of ZO-1, Claudin, and Occludin in the mouse colonic mucosa to varying degrees, and the expression level of tight junction proteins was significantly increased. P <0.05. Meanwhile, compared to the 20 mg / kg IBA treatment group, the 40 mg / kg IBA treatment group significantly restored the DSS-induced decrease in tight junction protein expression, and significantly increased the expression levels of ZO-1 and Claudin. P <0.05).

[0100] These results further demonstrate that IBA alleviates DSS-induced intestinal mucosal damage by restoring tight junction protein expression in the mouse colon, and that 40 mg / kg of IBA is more effective. The results and trends are largely consistent with those of RT-qPCR and ELISA.

[0101] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. The application of indole-3-butyric acid in the preparation of feed additives, characterized in that, The feed additive has any one or more of the following functions: (1) Alleviate oxidative stress and damage; (2) Improve growth performance; (3) Antidiarrheal; (4) Antioxidant; (5) Improve immune levels; (6) Regulates liver function; (7) Improve gut health; (8) Prevention and treatment of intestinal diseases.

2. The application according to claim 1, characterized in that, The improvement in growth performance includes increasing the daily weight gain of animals and / or increasing the apparent digestibility of nutrients.

3. The application according to claim 1, characterized in that, The antidiarrheal treatment includes reducing the diarrhea rate in animals, reducing the activity of diamine oxidase in animal serum, and / or reducing the content of D-lactic acid in animal serum.

4. The application according to claim 1, characterized in that, The intestinal diseases mentioned include colitis.

5. The application according to claim 1, characterized in that, The feed additives include pig feed additives.

6. A feed additive for improving animal gut health and preventing intestinal diseases, characterized in that, It contains indole-3-butyric acid.

7. The feed additive according to claim 6, characterized in that, The intestinal diseases mentioned include colitis.

8. The feed additive according to claim 6, characterized in that, The animals mentioned include pigs.

9. A feed for improving animal intestinal health and preventing intestinal diseases, characterized in that, It includes a basal diet and the feed additives described in any one of claims 6-8.

10. The feed according to claim 9, characterized in that, The animals mentioned include pigs.

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