Medicine for protecting intestinal health of chicken

Abstract

The invention relates to a pharmaceutical preparation for protecting the intestinal health of chickens, and belongs to the technical field of veterinary drugs. The core component is an oligopeptide composition (molecular weight lt: 1000 Da) prepared by enzymolysis of tilapia fish skin, solen meat and scapharca subcrenata meat according to a ratio of 4: 3: 3. The active oligopeptide with a synergistic effect is obtained through trypsin enzymolysis and an ultrafiltration purification process, so that proliferation of chicken small intestine epithelial cells can be remarkably promoted, and efficient protection on toxin damage can be realized. Animal experiments show that after 1 mg / kg of the oligopeptide composition is added into chicks fed with aflatoxin B1 polluted feed (100 [mu] g / kg), growth inhibition and diarrhea increase of the chicks caused by aflatoxin B1 can be remarkably reduced.

Description

Technical Field

[0001] The invention belongs to the technical field of veterinary medicines, and in particular relates to a medicine for protecting the intestinal health of chickens. Background Art

[0002] Intestinal health is a key factor in determining chicken production efficiency during the farming process. As a major component of the digestive system, the intestine is not only a key site for feed breakdown and nutrient absorption, but also plays a vital role in immune defense. A healthy intestine efficiently digests and absorbs nutrients, while also forming a first line of defense against pathogenic microorganisms, thereby safeguarding the body's overall health. Furthermore, intestinal health directly impacts a chicken's growth rate and feed conversion efficiency, making it a crucial foundation for efficient farming.

[0003] In actual farming environments, a variety of factors can threaten the intestinal health of chickens, including feed quality, environmental conditions, pathogen infection, and toxin exposure. These factors can lead to damage to small intestinal epithelial cells, bacterial imbalance, and decreased immune function, which can lead to indigestion, nutrient waste, and even disease transmission. Therefore, maintaining a healthy chicken intestine not only helps improve feed utilization but also significantly reduces disease incidence, lowers farming costs, and brings higher economic benefits to the farming industry. Summary of the Invention

[0004] The purpose of the present invention is to provide a drug for protecting the intestinal health of chickens, thereby promoting the development of the chicken intestine and alleviating the growth inhibition and diarrhea problems caused by aflatoxin B1.

[0005] To achieve the above object, the present invention provides the following technical solutions: First, the present invention provides a drug for protecting the intestinal health of chickens. The core active ingredient of the drug is an oligopeptide composition, which is prepared by enzymatic hydrolysis of tilapia skin, bamboo razor clam meat and hairy cockle meat. The weight ratio of the raw materials is tilapia skin: bamboo razor clam meat: hairy cockle meat = (1-6): (2-6): (1-7), and the molecular weight of the oligopeptide is less than 1000Da.

[0006] Preferably, the chicken intestinal health protection is to promote the proliferation of chicken small intestinal epithelial cells and reduce intestinal damage caused by aflatoxin B1.

[0007] Preferably, the method for preparing the oligopeptide composition comprises the following steps: (1) Raw material processing: clean the tilapia skin, bamboo clam meat and hairy clam meat, dry them to a moisture content of ≤8%, crush and sieve them, and mix them in a weight ratio of (1-6):(2-6):(1-7) to obtain a powder composition; (2) Enzymatic hydrolysis: add water at a solid-liquid ratio of 5:1, adjust the pH to 8, add 2% by weight of trypsin of the powder composition, and perform enzymatic hydrolysis at 37°C for 3 hours; (3) Inactivation and centrifugation: After inactivation, centrifuge and take the supernatant; (4) Ultrafiltration purification: After filtration through a 0.22 μm membrane, separation was performed using an ultrafiltration membrane with a molecular weight cutoff of 1000 Da to obtain an oligopeptide composition solution with a molecular weight of less than 1000 Da; (5) Concentration and drying: The oligopeptide composition solution is concentrated under reduced pressure to 1 / 5 of the original volume, and then freeze-dried to obtain the oligopeptide composition.

[0008] Preferably, the weight ratio of the raw materials is tilapia skin: razor clam meat: hairy clam meat = 4:3:3.

[0009] Secondly, the present invention provides an application of an oligopeptide composition in the preparation of a pharmaceutical preparation for promoting the growth of chicken small intestinal epithelial cells, characterized in that the oligopeptide composition is prepared from tilapia skin, bamboo razor clam meat and hairy cockle meat according to the preparation method described above.

[0010] Preferably, the weight ratio of the tilapia skin: razor clam meat: hairy cockle meat is 4:3:3.

[0011] Preferably, the concentration of the oligopeptide composition in the pharmaceutical preparation is 0.1-1.0 mg / mL.

[0012] In addition, the present invention provides an application of an oligopeptide composition in the preparation of a pharmaceutical preparation for protecting against growth inhibition and diarrhea in chickens caused by aflatoxin B1, characterized in that the oligopeptide composition is prepared from tilapia skin, bamboo clam meat and hairy clam meat according to the preparation method described above.

[0013] Preferably, the weight ratio of the tilapia skin: razor clam meat: hairy cockle meat is 4:3:3.

[0014] Preferably, the pharmaceutical preparation is added to a basic feed for use; The dosage of the pharmaceutical preparation added to 1 kg of the basic feed is 1 mg.

[0015] The beneficial effects of the present invention are: The present invention's oligopeptide composition (molecular weight <1000Da), prepared by enzymatic hydrolysis of tilapia skin, bamboo razor clam meat, and hairy cockle meat, significantly promoted the proliferation of chicken small intestinal epithelial cells and protected against aflatoxin B1-induced intestinal damage. In vitro experiments demonstrated that, at an optimal ratio (4:3:3), oligopeptide composition 1 achieved a cell proliferation rate of 194.64% at a concentration of 1.0 mg / mL, 1.33 times that of a single-source oligopeptide. Furthermore, the present oligopeptide composition, through a multi-target mechanism, increased the cell survival rate in an aflatoxin B1-damaged model to 80.38%, achieving a relative protection rate of 66.23%, significantly superior to single components or large-molecule peptides.

[0016] Animal experiments verified the practical application value of the composition. When 1 mg / kg of oligopeptide composition 1 was added to the diet of chicks fed 100 μg / kg of AFB1, the final body weight reached 146.27 g, and the diarrhea rate dropped to 3.14%. This not only exceeded the effect of a single-source oligopeptide, but also outperformed the control group to a certain extent. This shows that the oligopeptide composition of the present invention can effectively protect against growth inhibition and increased diarrhea caused by aflatoxin, while also promoting intestinal health and growth in chicks.

[0017] Therefore, the oligopeptide composition provided by the present invention can be used as an antibiotic-free functional additive to replace antibiotics for mycotoxin prevention and control, achieving the dual benefits of growth performance improvement and disease prevention at a low addition dosage (0.1-1.0 mg / mL), with significant economic and social benefits. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 The effects of different concentrations of oligopeptide compositions 1-4 on the proliferation of chicken small intestinal epithelial cells were demonstrated; Figure 1 Among them, (a) is the result of oligopeptide composition 1, (b) is the result of oligopeptide composition 2, (c) is the result of oligopeptide composition 3; (d) is the result of oligopeptide composition 4; Figure 2 The effects of different concentrations of tilapia skin oligopeptides, bamboo clam oligopeptides, hairy clam meat oligopeptides and peptide combinations on the proliferation of chicken small intestinal epithelial cells were demonstrated; Figure 2 Among them, (a) is the result of tilapia skin oligopeptide, (b) is the result of bamboo clam oligopeptide, (c) is the result of hairy clam oligopeptide, and (d) is the result of polypeptide combination; Figure 3 demonstrated the role of different oligopeptides in protecting chicken intestinal epithelial cell death induced by aflatoxin; Figure 4 The differences in terminal body weight among the different treatment groups are shown; Figure 5 The differences in diarrhea rates among the different treatment groups are shown. DETAILED DESCRIPTION

[0019] The present invention will be described and illustrated in more detail below through specific examples. However, it should be noted that these examples are only for illustrative purposes and do not constitute a limitation on the scope of protection of the present invention.

[0020] As a functional ingredient, oligopeptides play an important role in promoting intestinal health. However, existing technologies mostly focus on the application of single oligopeptides, and the mechanism of action and effect of such single ingredients are often relatively limited. To address this problem, the present invention proposes an innovative technical solution, namely, by scientifically proportioning and combining multiple preparation raw materials to prepare a mixed oligopeptide product. Through the synergistic effect between the various ingredients, a more efficient intestinal health promotion effect is achieved.

[0021] Example 1 Preparation of oligopeptide composition 1 (taking 100 g as an example) (1) Raw material processing Wash the tilapia skin, bamboo clam meat and hairy cockle meat, place them in a dryer and dry them until the moisture content is ≤8%; The dried tilapia skin, bamboo clam meat and hairy clam meat were crushed using a grinder, and then passed through an 80-mesh sieve to obtain tilapia skin powder, bamboo clam meat powder and hairy clam meat powder; (2) Enzymatic hydrolysis Mixing tilapia skin powder, bamboo clam meat powder, and hairy clam meat powder in a ratio of 4:3:3 to obtain a powder composition; 100 g of the powdered composition was mixed with deionized water at a solid-to-liquid ratio of 5:1, and stirred at 600 rpm for 30 minutes at room temperature to obtain a composition slurry; The homogenate of the composition was adjusted to 37° C. and pH 8, and trypsin with an enzyme activity of 2500 U / mg was added at an addition amount of 2% of the substrate, and enzymatic hydrolysis was performed for 3 hours; (3) Enzyme inactivation and centrifugation After enzymatic hydrolysis, the cells were rapidly heated to 95°C and kept warm for 15 minutes to inactivate trypsin. Allow to cool to room temperature, centrifuge at 8000 rpm for 15 minutes, and collect the supernatant; (4) Ultrafiltration separation The supernatant was preliminarily filtered using a 0.22 μm filter membrane to obtain a peptide composition solution; The combined peptide solution was filtered using an ultrafiltration membrane with a molecular cutoff of 1000 Da to obtain an oligopeptide composition solution; (5) Concentration and drying The oligopeptide composition solution was rotary evaporated under reduced pressure at 40° C. to 1 / 5 of the original volume to obtain a concentrated oligopeptide composition solution; The concentrated oligopeptide composition solution was placed in a freeze dryer for vacuum freeze drying to obtain oligopeptide composition 1.

[0022] Example 2 Preparation of oligopeptide composition 2 (1) Raw material processing Wash the tilapia skin, bamboo clam meat and hairy cockle meat, place them in a dryer and dry them until the moisture content is ≤8%; The dried tilapia skin, bamboo clam meat and hairy clam meat were crushed using a grinder, and then passed through an 80-mesh sieve to obtain tilapia skin powder, bamboo clam meat powder and hairy clam meat powder; (2) Enzymatic hydrolysis Mixing tilapia skin powder, bamboo clam meat powder, and hairy clam meat powder in a ratio of 6:3:1 to obtain a powder composition; 100 g of the powdered composition was mixed with deionized water at a solid-to-liquid ratio of 5:1, and stirred at 600 rpm for 30 minutes at room temperature to obtain a composition slurry; The homogenate of the composition was adjusted to 37° C. and pH 8, and trypsin with an enzyme activity of 2500 U / mg was added at an addition amount of 2% of the substrate, and enzymatic hydrolysis was performed for 3 hours; (3) Enzyme inactivation and centrifugation After enzymatic hydrolysis, the cells were rapidly heated to 95°C and kept warm for 15 minutes to inactivate trypsin. Allow to cool to room temperature, centrifuge at 8000 rpm for 15 minutes, and collect the supernatant; (4) Ultrafiltration separation The supernatant was preliminarily filtered using a 0.22 μm filter membrane to obtain a peptide composition solution; The combined peptide solution was filtered using an ultrafiltration membrane with a molecular cutoff of 1000 Da to obtain an oligopeptide composition solution; (5) Concentration and drying The oligopeptide composition solution was rotary evaporated under reduced pressure at 40° C. to 1 / 5 of the original volume to obtain a concentrated oligopeptide composition solution; The concentrated oligopeptide composition solution was placed in a freeze dryer for vacuum freeze drying to obtain oligopeptide composition 2.

[0023] Example 3 Preparation of oligopeptide composition 3 (1) Raw material processing Wash the tilapia skin, bamboo clam meat and hairy cockle meat, place them in a dryer and dry them until the moisture content is ≤8%; The dried tilapia skin, bamboo clam meat and hairy clam meat were crushed using a grinder, and then passed through an 80-mesh sieve to obtain tilapia skin powder, bamboo clam meat powder and hairy clam meat powder; (2) Enzymatic hydrolysis Mixing tilapia skin powder, bamboo clam meat powder, and hairy clam meat powder in a ratio of 2:6:2 to obtain a powder composition; 100 g of the powdered composition was mixed with deionized water at a solid-to-liquid ratio of 5:1, and stirred at 600 rpm for 30 minutes at room temperature to obtain a composition slurry; The homogenate of the composition was adjusted to 37° C. and pH 8, and trypsin with an enzyme activity of 2500 U / mg was added at an addition amount of 2% of the substrate, and enzymatic hydrolysis was performed for 3 hours; (3) Enzyme inactivation and centrifugation After enzymatic hydrolysis, the cells were rapidly heated to 95°C and kept warm for 15 minutes to inactivate trypsin. Allow to cool to room temperature, centrifuge at 8000 rpm for 15 minutes, and collect the supernatant; (4) Ultrafiltration separation The supernatant was preliminarily filtered using a 0.22 μm filter membrane to obtain a peptide composition solution; The combined peptide solution was filtered using an ultrafiltration membrane with a molecular cutoff of 1000 Da to obtain an oligopeptide composition solution; (5) Concentration and drying The oligopeptide composition solution was rotary evaporated under reduced pressure at 40° C. to 1 / 5 of the original volume to obtain a concentrated oligopeptide composition solution; The concentrated oligopeptide composition solution was placed in a freeze dryer for vacuum freeze drying to obtain oligopeptide composition 3.

[0024] Example 4 Preparation of oligopeptide composition 4 (1) Raw material processing Wash the tilapia skin, bamboo clam meat and hairy cockle meat, place them in a dryer and dry them until the moisture content is ≤8%; The dried tilapia skin, bamboo clam meat and hairy clam meat were crushed using a grinder, and then passed through an 80-mesh sieve to obtain tilapia skin powder, bamboo clam meat powder and hairy clam meat powder; (2) Enzymatic hydrolysis Mixing tilapia skin powder, bamboo clam meat powder, and hairy clam meat powder in a ratio of 1:2:7 to obtain a powder composition; 100 g of the powdered composition was mixed with deionized water at a solid-to-liquid ratio of 5:1, and stirred at 600 rpm for 30 minutes at room temperature to obtain a composition slurry; The homogenate of the composition was adjusted to 37° C. and pH 8, and trypsin with an enzyme activity of 2500 U / mg was added at an addition amount of 2% of the substrate, and enzymatic hydrolysis was performed for 3 hours; (3) Enzyme inactivation and centrifugation After enzymatic hydrolysis, the cells were rapidly heated to 95°C and kept warm for 15 minutes to inactivate trypsin. Allow to cool to room temperature, centrifuge at 8000 rpm for 15 minutes, and collect the supernatant; (4) Ultrafiltration separation The supernatant was preliminarily filtered using a 0.22 μm filter membrane to obtain an oligopeptide composition solution; The combined peptide solution was filtered using an ultrafiltration membrane with a molecular cutoff of 1000 Da to obtain an oligopeptide composition solution; (5) Concentration and drying The oligopeptide composition solution was rotary evaporated under reduced pressure at 40° C. to 1 / 5 of the original volume to obtain a concentrated oligopeptide composition solution; The concentrated oligopeptide composition solution was placed in a freeze dryer for vacuum freeze drying to obtain oligopeptide composition 4.

[0025] Comparative Example 1 Tilapia skin oligopeptides were prepared using only tilapia skin as the raw material, using the same preparation method as in Example 1.

[0026] Comparative Example 2 Only bamboo clam meat was used as the raw material, and the same preparation method as in Example 1 was used to prepare bamboo clam oligopeptides.

[0027] Comparative Example 3 The raw material used was only razor clam meat, and the razor clam oligopeptide was prepared using the same preparation method as in Example 1.

[0028] Comparative Example 4 The raw materials were prepared in the same manner as in Example 1. After separation by ultrafiltration membrane, polypeptides with a molecular weight greater than 1000 Da were collected to prepare a polypeptide composition.

[0029] Example 5 Effects of different oligopeptides on the proliferation of chicken small intestinal epithelial cells (1) Chicken intestinal epithelial cells (CSIEC purchased from Shanghai Qingqi Biotechnology) were cultured in DMEM / F12 medium containing 10% fetal bovine serum at 37°C and 5% CO2 until the logarithmic growth phase.

[0030] (2) After digestion, the cells were inoculated into 96-well plates, with 1×10 cells per well. 4 cells; (3) After the cells are completely attached, they are treated according to the following groups: Experimental groups: Blank control group: containing only basal culture medium.

[0031] Groups of Examples 1-4: oligopeptide compositions 1-4 were added at final concentrations of 0.1 mg / mL, 0.5 mg / mL, and 1.0 mg / mL, respectively.

[0032] Comparative Examples 1-4: Tilapia oligopeptide (Comparative Example 1), bamboo clam oligopeptide (Comparative Example 2), hairy clam oligopeptide (Comparative Example 3) and polypeptide combination (Comparative Example 4) were added at the same concentrations respectively; Five replicate wells were set up for each treatment group.

[0033] After culturing for 24 h, the absorbance of each group was measured using CCK-8, and the cell proliferation rate was calculated based on the control group.

[0034] The experimental results are as follows Figure 1 and Table 1 (Oligopeptide Compositions 1-4) and Figure 2 and as shown in Table 2 (single-source oligopeptide and polypeptide compositions).

[0035] Table 1 Effects of oligopeptide compositions 1-4 on the proliferation of chicken small intestinal epithelial cells

[0036] Table 2 Effects of tilapia oligopeptides, bamboo clam oligopeptides, hairy clam oligopeptides and polypeptide combination on the proliferation of chicken small intestinal epithelial cells

[0037] From Table 1-Table 2 and Figure 1 and Figure 2 The results show that, with the same total amount of raw materials, at a low concentration of 0.1 mg / mL, the cell proliferation rate of oligopeptide composition 1 was 130.63%, significantly higher than the maximum value of a single-source oligopeptide in Table 2 (razor clam oligopeptide: 111.77%). This indicates that the oligopeptide composition can exert a significant pro-proliferation effect at a low dose, while the effect of a single-source oligopeptide is relatively limited.

[0038] At a higher concentration of 1 mg / mL, the cell proliferation rate of oligopeptide composition 1 (194.64%) was 1.33 times the maximum value of a single-source oligopeptide (razor clam oligopeptide: 146.19%), further verifying the synergistic effect of the oligopeptide composition.

[0039] Secondly, it can be seen that there are significant differences in the cell proliferation rates of different oligopeptide compositions (1-4) in Table 1, among which oligopeptide composition 1 (tilapia skin: bamboo razor clam meat: hairy cockle meat = 4:3:3) has the best effect at all concentrations: At 0.1 mg / mL, oligopeptide composition 1 (130.63%) was significantly higher than other compositions (such as oligopeptide composition 2: 118.51%; oligopeptide composition 3: 117.36%; oligopeptide composition 4: 111.83%).

[0040] At 1 mg / mL, the cell proliferation rate of oligopeptide composition 1 (194.64%) was also much higher than that of other compositions (such as oligopeptide composition 2: 163.58%; oligopeptide composition 3: 167.28%; oligopeptide composition 4: 156.27%).

[0041] This shows that the ratio of raw materials has a significant impact on the functionality of the oligopeptide composition. The ratio of oligopeptide composition 1 (4:3:3) is the best, maximizing the synergistic effect between the components. Although the effect of oligopeptide composition 4 is higher than that of single-source oligopeptides, the improvement is smaller.

[0042] In addition, the cell proliferation rate of the peptide composition (molecular weight > 1000 Da) in Table 2 was significantly lower than that of oligopeptide composition 1 (<1000 Da) at all concentrations, indicating the necessity of molecular weight screening. It also indicates that simply combining tilapia skin, bamboo razor clam meat, and hairy cockle meat without controlling the molecular weight (>1000 Da) does not achieve a synergistic effect in promoting the proliferation of chicken small intestinal epithelial cells.

[0043] Example 6 Effects of different oligopeptides on protecting chicken small intestinal epithelial cell death induced by aflatoxin B1 (1) Chicken small intestinal epithelial cells were cultured in DMEM / F12 medium containing 10% fetal bovine serum at 37°C and 5% CO2 until the logarithmic growth phase.

[0044] (2) After digestion, the cells were inoculated into 96-well plates, with 1×10 cells per well. 4 cells; (3) After the cells are cultured to a cell density of more than 90%, the original culture medium is removed and the cells are treated according to the following groups: Control group: After treatment with DMEM / F12 serum-free medium for 6 hours, the medium was replaced with DMEM / F12 medium containing 10% fetal bovine serum and cultured for another 24 hours; Injury group: After treatment with DMEM / F12 serum-free medium for 6 h, the medium was replaced with DMEM / F12 medium containing 300 μmol / L aflatoxin B1 and 10% fetal bovine serum and cultured for another 24 h; Protection group 1: After adding DMEM / F12 serum-free medium containing oligopeptide composition 1 (1 mg / mL, the concentration of subsequent protection groups was the same) for 6 hours, the medium was replaced with DMEM / F12 medium containing 300 μmol / L aflatoxin B1 and 10% fetal bovine serum and cultured for another 24 hours; Protection group 2: After treatment with DMEM / F12 serum-free medium containing oligopeptide composition 2 for 6 hours, the medium was replaced with DMEM / F12 medium containing 300 μmol / L aflatoxin B1 and 10% fetal bovine serum and cultured for another 24 hours; Protection group 3: After treatment with DMEM / F12 serum-free medium containing oligopeptide composition 3 for 6 hours, the medium was replaced with DMEM / F12 medium containing 300 μmol / L aflatoxin B1 and 10% fetal bovine serum and cultured for another 24 hours; Protection group 4: After treatment with DMEM / F12 serum-free medium containing oligopeptide composition 4 for 6 hours, the medium was replaced with DMEM / F12 medium containing 300 μmol / L aflatoxin B1 and 10% fetal bovine serum and cultured for another 24 hours; Protection group 5: After adding DMEM / F12 serum-free medium containing tilapia oligopeptide for 6 hours, the medium was replaced with DMEM / F12 medium containing 300 μmol / L aflatoxin B1 and 10% fetal bovine serum and cultured for another 24 hours; Protection group 6: After adding DMEM / F12 serum-free medium containing bamboo clam oligopeptides for 6 hours, the medium was replaced with DMEM / F12 medium containing 300 μmol / L aflatoxin B1 and 10% fetal bovine serum and cultured for another 24 hours; Protection group 7: After treatment with DMEM / F12 serum-free medium containing razor clams oligopeptide for 6 hours, the medium was replaced with DMEM / F12 medium containing 300 μmol / L aflatoxin B1 and 10% fetal bovine serum and cultured for another 24 hours; Protection group 8: After adding the peptide composition-containing DMEM / F12 serum-free medium for 6 hours, the medium was replaced with DMEM / F12 medium containing 300 μmol / L aflatoxin B1 and 10% fetal bovine serum and cultured for another 24 hours; Five replicate wells were set up for each treatment group; After the culture was completed, the absorbance of each well was measured using CCK-8 reagent, and the cell survival rate of each group was calculated based on the control group. The results are shown in Table 3 and Figure 3 shown.

[0045] Table 3 Effects of different oligopeptides on protecting chicken intestinal epithelial cell death caused by aflatoxin

[0046] From Table 3 and Figure 3 The results show that the cell survival rate in the injury group is only 41.15%, which is much lower than 100% in the control group. This shows that AFB1 can effectively induce toxic reactions in chicken small intestinal epithelial cells at a concentration of 300 μmol / L, causing obvious cell death, indicating that the present invention has constructed a stable cell injury model.

[0047] Results from protection groups 1-4 showed that protection group 1 (oligopeptide composition 1) achieved a cell survival rate of 80.38% and a relative damage protection rate of 66.23%, significantly superior to the other protection groups, demonstrating the composition's outstanding ability to mitigate AFB1 toxicity. This suggests that, under the specific ratio of tilapia skin: bamboo razor clam meat: hairy cockle meat = 4:3:3, the oligopeptides may have a significant synergistic effect, effectively enhancing cellular antioxidant capacity and membrane stability, and inhibiting toxin-induced apoptosis.

[0048] In contrast, Protective Group 4 (Oligopeptide Combination 4), although also composed of three ingredients but in a different ratio (3:2:5), achieved a cell survival rate of 57.44% and a relative protection rate of 27.68%. While slightly higher than the single-source oligopeptides, the improvement was limited. This result suggests that not all combinations of ingredients can produce a significant synergistic protective effect; only when the ingredients are scientifically and rationally proportioned can the synergistic effects of the oligopeptides be fully realized.

[0049] Results from protection groups 5-7 showed that the cell survival rates of the three were 50.94%, 54.68%, and 50.22%, respectively, only slightly higher than the injury group (41.15%). This suggests that while oligopeptides from a single source have some protective effects, their functions are relatively limited and they are unable to activate multiple cellular defense mechanisms. Therefore, their overall protective effect is limited and they are unable to cope with complex toxin challenges.

[0050] In protection group 8, the survival rate of the polypeptide composition was low and could not exert sufficient protection effect.

[0051] Example 7 Protective effect of oligopeptide combination on aflatoxin damage in broiler chickens (1) Two hundred one-day-old AA broiler chicks were randomly assigned to four treatment groups, with five replicates per treatment and ten chicks per replicate. The chicks were fed according to the following groups: Control group: fed with basal diet; Damage group: fed with basal diet and 100 μg / kg aflatoxin B1; Protection group 1: fed with basal diet, 100 μg / kg aflatoxin B1 and 1 mg / kg bamboo clam oligopeptide; Protection group 2: fed with basal diet, 100 μg / kg aflatoxin B1 and 1 mg / kg oligopeptide composition 1; The basal diet consisted of: 52.07% corn, 35.50% soybean meal, 5% fish meal, 4% soybean oil, 1% calcium hydrogen phosphate, 1% rock powder, 0.3% salt, 0.04% lysine, 0.09% methionine, and 1% premix; (2) After 21 days of feeding, the final weight and diarrhea rate of the chicks were tested. The diarrhea rate = total diarrhea days / (total number of chickens × number of test days).

[0052] The results are shown in Table 4 and Figure 4 and Figure 5 As shown: Table 4 Protective effect of oligopeptide composition on aflatoxin damage in broiler chickens

[0053] From Table 4, Figure 4 and Figure 5 The results showed that compared with the control group, the final body weight of the chickens in the damaged group fed with aflatoxin B1 was significantly reduced (134.49±4.99 g vs. 144.26±3.36 g), and the diarrhea rate was significantly increased (6.85% vs. 3.39%), indicating that aflatoxin B1 successfully induced a growth inhibition and intestinal health damage model in broiler chickens.

[0054] Further comparison of the protection group and the injury group revealed: The final body weight of protection group 1 (intervention with bamboo clam oligopeptides) was 5.44 g higher than that of the injury group (139.93±3.20 g vs. 134.49±4.99 g), and the diarrhea rate decreased to 5.09%, indicating that bamboo clam oligopeptides have a certain alleviating effect on AFB1 toxicity, but the overall effect is relatively limited.

[0055] The final body weight of Protected Group 2 (intervention with Oligopeptide Composition 1) reached 146.27±4.76 g, significantly higher than both the Injured Group and Protected Group 1, and even exceeding the Control Group, indicating that the oligopeptide composition also promotes growth. Furthermore, the diarrhea rate in the Protected Group decreased to 3.14%, significantly lower than that in Protected Group 1 and slightly lower than that in the Control Group. These results suggest that Oligopeptide Composition 1, through the synergistic effects of tilapia skin oligopeptides, bamboo razor clam meat oligopeptides, and hairy clam meat oligopeptides, can reduce intestinal damage caused by aflatoxin B1 while promoting growth in chicks.

Claims

1. A drug for protecting the intestinal health of chickens, characterized in that: The core active ingredient of the drug is an oligopeptide composition, which is prepared by enzymatic hydrolysis of tilapia skin, bamboo razor clam meat and hairy cockle meat. The weight ratio of the raw materials is tilapia skin: bamboo razor clam meat: hairy cockle meat = (1-6): (2-6): (1-7), and the molecular weight of the oligopeptide is less than 1000Da.

2. The drug according to claim 1, characterized in that The chicken intestinal health protection is to promote the proliferation of chicken small intestinal epithelial cells and reduce the intestinal damage caused by aflatoxin B1.

3. The drug according to claim 2, characterized in that The preparation method of the oligopeptide composition comprises the following steps: (1) Raw material processing: clean the tilapia skin, bamboo clam meat and hairy clam meat, dry them to a moisture content of ≤8%, crush and sieve them, and mix them in a weight ratio of (1-6):(2-6):(1-7) to obtain a powder composition; (2) Enzymatic hydrolysis: add water at a solid-liquid ratio of 5:1, adjust the pH to 8, add trypsin at 2% by weight of the powder composition, and perform enzymatic hydrolysis at 37°C for 3 hours; (3) Inactivation and centrifugation: After inactivation, centrifuge and take the supernatant; (4) Ultrafiltration purification: After filtration through a 0.22 μm membrane, separation was performed using an ultrafiltration membrane with a molecular weight cutoff of 1000 Da to obtain an oligopeptide composition solution with a molecular weight of less than 1000 Da; (5) Concentration and drying: The oligopeptide composition solution is concentrated under reduced pressure to 1 / 5 of the original volume, and then freeze-dried to obtain the oligopeptide composition.

4. The drug according to claim 3, characterized in that The weight ratio of the raw materials is tilapia skin: razor clam meat: hairy clam meat = 4:3:

3.

5. Use of an oligopeptide composition in the preparation of a pharmaceutical preparation for promoting the growth of chicken small intestinal epithelial cells, characterized in that: The oligopeptide composition is prepared from tilapia skin, bamboo razor clam meat and hairy clam meat according to the preparation method described in claim 3.

6. The use according to claim 5, characterized in that The weight ratio of the tilapia skin, bamboo clam meat and hairy clam meat is 4:3:

3.

7. The use according to claim 6, characterized in that In the pharmaceutical preparation, the concentration of the oligopeptide composition is 0.1-1.0 mg / mL.

8. Use of an oligopeptide composition in the preparation of a pharmaceutical preparation for protecting chickens from growth inhibition and diarrhea caused by aflatoxin B1, characterized in that: The oligopeptide composition is prepared from tilapia skin, bamboo razor clam meat and hairy clam meat according to the preparation method described in claim 3.

9. The use according to claim 8, characterized in that The weight ratio of the tilapia skin, bamboo clam meat and hairy clam meat is 4:3:

3.

10. The use according to claim 9, characterized in that The pharmaceutical preparation is added to basic feed for use; The dosage of the pharmaceutical preparation added to 1 kg of the basic feed is 1 mg.

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