A black-bone chicken liver hydrolyzed peptide capable of improving ulcerative colitis, and its preparation method and application

Abstract

The present invention discloses a black chicken liver hydrolyzed peptide that can be used to improve ulcerative colitis, and its preparation method and application, which belong to the field of biomedicine technology. The preparation method of the black chicken liver hydrolyzed peptide comprises the following steps: making black chicken liver into minced meat and mixing it with water, heating the obtained liquid material, adding alkaline protease for enzymatic hydrolysis, inactivating the enzyme and collecting the supernatant by centrifugation, and freeze-drying to obtain the black chicken liver hydrolyzed peptide. The present invention adopts formaldehyde titration to detect the degree of hydrolysis, adopts SDS-PAGE electrophoresis and Tricine-SDS-PAGE electrophoresis to detect the molecular weight, performs sequence analysis by LC-MS / MS, uses a database to predict the biological function of peptide segments, detects the antioxidant and blood pressure lowering ability of black chicken liver peptide by in vitro experiments, and detects the effect of black chicken liver peptide in improving colitis by mouse experiments. The results show that the black chicken liver peptide prepared by the present invention has the advantages of antioxidant, blood pressure lowering and improvement of ulcerative colitis.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedicine, and in particular to a black-bone chicken liver hydrolyzed peptide capable of improving ulcerative colitis, and a preparation method and application thereof. Background Art

[0002] Ulcerative colitis (UC) is a recurrent inflammatory bowel disease. As one of the main types of inflammatory bowel disease, UC is characterized by weight loss, abdominal pain, diarrhea, and bloody stools. Conventional UC intervention drugs include corticosteroids, 5-aminosalicylic acid, immunosuppressants, and antibiotics, but their effectiveness is limited and accompanied by a series of side effects. Long-term toxicity also limits their efficacy. Therefore, it is crucial to explore safe and effective ways to prevent or improve UC.

[0003] Chicken liver, an edible byproduct of chicken slaughter, is rich in protein, calcium, phosphorus, iron, zinc, vitamin A, and B vitamins. However, fresh chicken liver is inexpensive and has a distinct fishy smell, making it rarely consumed directly. Most is used as animal feed or even discarded. Only a small amount is further processed into braised liver and chicken liver pâté, resulting in a significant waste of high-quality protein. Therefore, increasing the value of chicken liver resources is of great practical significance.

[0004] Chinese patent publication CN118685483A discloses a method for preparing a black chicken peptide with blood-tonifying properties. The resulting black chicken peptide boasts high oligopeptide content, a mild bitter, astringent, and salty taste, excellent antioxidant activity, high purity, and a potent blood-tonifying effect. Current research on black chicken peptides primarily focuses on their antioxidant, immune-regulating, and blood-tonifying properties. However, there is a lack of research on the preparation of black chicken liver peptides and their application in improving colitis. Summary of the Invention

[0005] The present invention aims to provide a hydrolyzed black-bone chicken liver peptide that can be used to improve ulcerative colitis, as well as its preparation method and application, to address the problems of the above-mentioned prior art. The hydrolyzed black-bone chicken liver peptide prepared by the present invention has the advantages of anti-oxidation, lowering blood pressure, and improving ulcerative colitis.

[0006] To achieve the above object, the present invention provides the following solutions:

[0007] Technical solution 1: A method for preparing hydrolyzed peptides from black-bone chicken liver, comprising the step of enzymatically hydrolyzing the black-bone chicken liver using alkaline protease.

[0008] The black-bone chicken comes from the Luoyeyang black-bone chicken in Shaanxi.

[0009] Furthermore, the preparation method comprises the following steps: preparing black-bone chicken liver into minced meat and mixing it with water, heating the obtained liquid, adding alkaline protease for enzymolysis, inactivating the enzyme and collecting the supernatant by centrifugation, and freeze-drying to obtain the black-bone chicken liver hydrolyzed peptide.

[0010] The heating temperature is 55°C.

[0011] Furthermore, the black-bone chicken liver is made into minced meat and mixed with water, and the minced meat is adjusted to a mass concentration of 30% by adding water.

[0012] Furthermore, the addition amount of the alkaline protease is 1% of the mass of the minced meat in the liquid material.

[0013] Furthermore, the enzymatic hydrolysis is carried out at 55° C. for 1 hour.

[0014] Furthermore, the enzyme is inactivated at 90° C. for 15 minutes.

[0015] Furthermore, the centrifugation is carried out at 8000 r / min at 4° C. for 10 min.

[0016] Technical solution 2: The black-bone chicken liver hydrolyzed peptide prepared by the preparation method.

[0017] Technical solution three: Use of the black-bone chicken liver hydrolyzed peptide in the preparation of a product having antioxidant activity and blood pressure lowering effects.

[0018] Technical solution 4: Use of the black-bone chicken liver hydrolyzed peptide in the preparation of a product having the effect of improving ulcerative colitis.

[0019] The products include functional foods and medicines.

[0020] The present invention discloses the following technical effects:

[0021] The invention obtains black-bone chicken liver hydrolyzed peptides (black-bone chicken liver peptides) by enzymatically hydrolyzing Lueyang black-bone chicken livers, detects the degree of hydrolysis by formaldehyde titration, detects the molecular weight by SDS-PAGE electrophoresis and Tricine-SDS-PAGE electrophoresis, performs sequence analysis by LC-MS / MS, predicts the biological functions of peptide segments by using a database, detects the antioxidant and blood pressure-lowering abilities of the black-bone chicken liver peptides by in vitro experiments, and detects the effect of the black-bone chicken liver peptides on improving colitis by mouse experiments.

[0022] The results show that the black chicken liver peptide prepared by the present invention is light yellow, with a crispy and porous surface; the present invention adopts formaldehyde titration method for detection, and the results show that the black chicken liver has reached a high degree of hydrolysis in 1 hour, and the amino nitrogen content in the black chicken liver peptide obtained by hydrolysis for 1 hour reaches 70.43% of that obtained by hydrolysis for 4 hours; the present invention adopts SDS-PAGE electrophoresis and Tricine-SDS-PAGE electrophoresis to detect that the molecular weight of black chicken liver is between 20-120kDa, and the molecular weight of black chicken liver peptide is between 4.1-6.5kDa, and as the hydrolysis time is prolonged, the molecular weight range of the peptide segment decreases; the present invention identifies and analyzes the sequence of black chicken liver peptide based on LC-MS / MS mass spectrometry detection method and finds that the number of peptide segments after hydrolysis of black chicken liver is 6637, and analyzes the amino acid composition ratio of black chicken liver peptide and finds that black chicken liver peptide contains 20 amino acids, including 9 essential amino acids, accounting for 42.7%, and has high nutritional value to the human body; Ming also found through comparison and prediction of the BioPepDB database that black chicken liver peptide has 487 peptide segments with antibacterial activity, 156 peptide segments with antihypertensive activity, and 2 peptide segments with anticancer activity; the antioxidant activity test found that compared with black chicken liver, the black chicken liver peptide prepared by the present invention has enhanced scavenging ability for DPPH free radicals and ABTS free radicals, and the black chicken liver peptide has a scavenging rate of 80.92% for DPPH free radicals and a scavenging rate of 95.83% for ABTS free radicals; the present invention also found through antihypertensive ability test that black chicken liver peptide has ACE2 inhibitory activity, with an inhibition rate of 12.35%; the results of mouse colon length, DAI score and colon section showed that black chicken liver peptide has the effect of alleviating ulcerative colitis; the colon section results of the present invention showed that black chicken liver peptide can significantly reduce the colon tissue damage of mice induced by sodium dextran sulfate; serum inflammatory factor detection showed that black chicken liver peptide has an improving effect on the level of intestinal inflammation induced by DSS. The above results show that the black chicken liver peptide prepared by the present invention has the advantages of anti-oxidation, lowering blood pressure and improving ulcerative colitis. Furthermore, using it to prepare products for preventing or improving ulcerative colitis is of great significance for improving the living conditions of patients with ulcerative colitis. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0024] Figure 1 This is the surface condition of black chicken liver peptide;

[0025] Figure 2 The amino acid composition of black-bone chicken liver peptide;

[0026] Figure 3 Comparison of the antioxidant capacity (DPPH free radical and ABTS free radical) of black-bone chicken liver and black-bone chicken liver peptide, where BL is black-bone chicken liver; BLP is black-bone chicken liver peptide;

[0027] Figure 4 The colon length statistics of mice in each group;

[0028] Figure 5 DAI scores of mice in each group;

[0029] Figure 6 HE staining of mouse colon tissue (50 μm), where NC is the normal group (A); DSS is the enteritis model group (B); ASA is the group administered with 40 mg / kg 5-aminosalicylic acid by gavage (C); BLPL is the group administered with 100 mg / kg black chicken liver peptide by gavage (D); BLPH is the group administered with 300 mg / kg black chicken liver peptide by gavage (E);

[0030] Figure 7 The expression of various inflammatory factors in mouse serum, among which A is IL-6; B is IL-1β; C is TNF-α; and D is IL-10. DETAILED DESCRIPTION

[0031] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0032] It should be understood that the terms described herein are intended only to describe particular embodiments and are not intended to limit the present invention. In addition, for numerical ranges herein, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. The intermediate value within any stated value or stated range, and each smaller range between any other stated value or intermediate value within the stated range, is also encompassed within the present invention. The upper and lower limits of these smaller ranges may be independently included or excluded within the scope.

[0033] Unless otherwise indicated, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein may also be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of any conflict with any incorporated document, the contents of this specification shall prevail.

[0034] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments described herein without departing from the scope or spirit of the invention. Other embodiments will be apparent to those skilled in the art from the description of the invention. The description and examples are intended to be illustrative only.

[0035] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0036] The present invention is further described below by way of specific examples. Unless otherwise specified, the various chemical reagents used in the examples of the present invention were obtained through conventional commercial channels.

[0037] Alkaline protease was purchased from Dongheng Huadao Biotechnology Co., Ltd. with an enzyme activity of 200,000 U / g;

[0038] Example 1

[0039] 1. Preparation of Black Chicken Liver Hydrolyzed Peptide (Black Chicken Liver Peptide)

[0040] Use a meat grinder to grind black chicken liver (from Lueyang black chicken in Lueyang County, Shaanxi Province) into minced meat, add water to adjust the mass concentration to 30%, heat the liquid to 55℃, add 1% alkaline protease of the minced meat mass, natural pH, enzymolysis at 55℃ for 1h, inactivate the enzyme at 90℃ for 15min, centrifuge at 8000r / min for 10min at 4℃, collect the supernatant, filter and retain the filtrate, vacuum freeze-dry the filtrate to obtain black chicken liver peptide. The obtained black chicken liver peptide is light yellow, with a crispy and porous surface ( Figure 1 ).

[0041] 2. Determination of physical and chemical properties

[0042] 2.1 Detection of hydrolysis degree of black-bone chicken liver by formaldehyde potentiometric titration

[0043] Following the method in step 1, enzymatic hydrolyses (i.e., filtrates) of black-bone chicken liver peptides were prepared after 1 hour and 4 hours of enzymatic hydrolysis, respectively. 15 mL of each enzymatic hydrolysate was taken, the pH adjusted to 8.2 with 0.05 mol NaOH standard solution, 5 mL of formaldehyde was added, and the solution was titrated to pH 9.2 with 0.05 mol NaOH standard solution. The sodium hydroxide consumption (V1) was recorded. Water was used as a blank control, and the sodium hydroxide consumption (V2) was recorded. The mass fraction of amino nitrogen was calculated using the following formula: X (mg / g) = [(V1-V2) × C × 14 × (50 / 15)] / 0.2, where X is the mass fraction of amino nitrogen; C is the mass molar concentration of NaOH; and V1 and V2 are the volumes of sodium hydroxide consumed. The results showed that black-bone chicken liver had reached a high degree of hydrolysis after 1 hour, with the amino nitrogen content of the black-bone chicken liver peptide obtained after 1 hour of hydrolysis reaching 70.43% of that after 4 hours of hydrolysis.

[0044] 2.2 Molecular weight detection

[0045] SDS-PAGE electrophoresis: Mix black chicken liver (BL) and black chicken liver peptide (BLP) with 5× protein loading buffer at a ratio of 4:1. Boil for 10 minutes and perform SDS-PAGE electrophoresis. Prepare the SDS-PAGE gel according to Table 1, loading 4 μL of marker and 6 μL of sample. Initially, run the gel at a constant voltage of 80 V. Once the gel is separated, adjust the voltage to 120 V until the end of the electrophoresis. Destain with Coomassie Brilliant Blue until the desired band is visible.

[0046] Table 1 SDS-PAGE gel formula

[0047]

[0048]

[0049] Tricine-SDS-PAGE electrophoresis: Mix black chicken liver peptide with 2× protein loading buffer in a 1:1 ratio and boil for 10 minutes for Tricine-SDS-PAGE electrophoresis. Prepare the Tricine-SDS-PAGE gel according to Table 2, with a marker loading volume of 5 μL and a sample loading volume of 10 μL. Run the gel at a constant current of 20 mA, then adjust to 30 mA at the separation gel until the gel is separated. Destain with Coomassie Brilliant Blue until the target band is visible.

[0050] Preparation of black-bone chicken liver (BL): Use a meat grinder to grind the black-bone chicken liver into minced meat, add water to adjust the mass concentration to 30%, heat the liquid to 55°C, natural pH, treat at 55°C for 1 hour, treat at 90°C for 15 minutes, centrifuge at 8000 rpm for 10 minutes at 4°C, collect the supernatant, filter and retain the filtrate, and vacuum freeze-dry the filtrate to obtain black-bone chicken liver.

[0051] Electrophoresis detection steps of black chicken liver: (1) Mix black chicken liver with 5× protein loading buffer at a ratio of 4:1, boil for 10 minutes and perform SDS-PAGE electrophoresis detection. The sample volume is 6 μL. First, use 80V constant voltage, and adjust the voltage to 120V when the gel is separated and run until the end. Use Coomassie Brilliant Blue to decolorize until the target band appears. (2) Mix black chicken liver with 2× protein loading buffer at a ratio of 1:1, boil for 10 minutes and perform Tricine-SDS-PAGE electrophoresis detection. The sample volume is 10 μL. First, use 20mA constant current, and adjust the voltage to 30mA when the gel is separated and run until the end. Use Coomassie Brilliant Blue to decolorize until the target band appears.

[0052] Table 2 Tricine-SDS-PAGE gel formula

[0053]

[0054] The results showed that the molecular weight of black-bone chicken liver protein detected by SDS-PAGE electrophoresis and Tricine-SDS-PAGE electrophoresis was between 20-120 kDa, and the molecular weight of black-bone chicken liver peptide was between 4.1-6.5 kDa. As the hydrolysis time increased, the molecular weight range of the peptide segment decreased.

[0055] 3. Statistics of peptide identification results and analysis of amino acid composition ratio

[0056] The analytical column was a 75 μm ID × 25 cm NanoViper C18 1.9 μm, 100 Å column. Mobile phase A was 0.1% FA, and mobile phase B was 0.1% FA, 80% ACN. The flow rate was 600 nL / min, and the analysis time for each component was 35 min. Specific chromatographic conditions are shown in Table 3.

[0057] Table 3 Liquid chromatography conditions

[0058]

[0059] The full mass spectrometry scan range was 100-1500 m / z, the primary mass spectrometry resolution was set to 60,000, AGC was set to Custom, and MaximumIT was set to Custom. The secondary mass spectrometry resolution was set to Resolution: 15,000, AGC was set to Custom, MaximumIT was set to Custom, Cycletime was set to 2s, and the peptide fragmentation collision energy was set to 32. The raw mass spectrometry data (raw) was generated. The raw mass spectrometry files were searched in the software database to obtain the identification results and the number of peptides for each sample. Each amino acid has different physicochemical properties and corresponding functions. For peptides, the amino acid sequence composition determines the physicochemical properties and functions of the peptides.

[0060] The results showed that the sequence identification and analysis of black chicken liver peptides based on LC-MS / MS mass spectrometry revealed that the number of peptides after hydrolysis of black chicken liver was 6637. Figure 2 ) found that black chicken liver peptide contains 20 kinds of amino acids, including 9 essential amino acids, accounting for 42.7%, and has high nutritional value to the human body.

[0061] 4. Peptide Function Prediction

[0062] Use the BioPepDB database to predict peptide functions. The peptide sequences obtained from peptide analysis are used for function prediction in the BioPepDB database. Click Get Result in the Prediction section to predict the function of the corresponding peptide.

[0063] The results showed that through comparison and prediction of the BioPepDB database, black chicken liver peptide was found to have 487 peptides with antibacterial activity, 156 peptides with antihypertensive activity, and 2 peptides with anticancer activity.

[0064] 5. Antioxidant Activity Detection of Black-bone Chicken Liver and Black-bone Chicken Liver Peptide

[0065] For details on the preparation of black-bone chicken liver (BL), please refer to "2.2 Molecular Weight Detection"; for details on the preparation of black-bone chicken liver peptide (BLP), please refer to "1. Preparation of Black-bone Chicken Liver Hydrolyzed Peptide (Black-bone Chicken Liver Peptide)".

[0066] 5.1 DPPH free radical scavenging rate determination: The DPPH free radical scavenging rate of black chicken liver and filtered black chicken liver hydrolyzed peptide filtrate was detected using the Shanghai Biotech Total Antioxidant Capacity (T-AOC) Detection Kit (DPPH method) microassay kit. Preheat the microplate reader for 30 minutes and adjust the wavelength to 515nm. React in a 1.5mL centrifuge tube, and the sample addition method is shown in Table 4. After the addition is completed, mix thoroughly, react at room temperature in the dark for 20 minutes, and take 200μL to a 96-well plate to measure the absorbance at 515nm. The calculation formula is as follows: DPPH free radical scavenging rate (%) = (A 空白 -A 测定 ) / A 空白 ×100%.

[0067] Table 4 DPPH method sample addition method

[0068]

[0069]

[0070] 5.2 ABTS free radical scavenging rate determination

[0071] The total antioxidant capacity (T-AOC) detection kit (ABTS method) of Shanghai Biotech was used to detect the ABTS free radical scavenging rate of black chicken liver and filtered black chicken liver hydrolyzed peptide filtrate. The microplate reader was preheated for 30 minutes and the wavelength was adjusted to 734nm. The reaction was carried out in a PCR tube and the sample addition method was shown in Table 5. After the sample addition was completed, the mixture was thoroughly mixed, the reaction was allowed to stand for 20 minutes, and the absorbance at 734nm was measured by transferring to a 96-well plate. The calculation formula is as follows: ABTS free radical scavenging rate (%) = (A 空白 -A 测定 ) / A 空白 ×100%.

[0072] Table 5 ABTS method sample addition method

[0073]

[0074] Antioxidant activity tests showed that compared with black chicken liver, black chicken liver peptide had enhanced scavenging ability for DPPH free radicals and ABTS free radicals. The scavenging rate of black chicken liver peptide for DPPH free radicals was 80.92%, and the scavenging rate for ABTS free radicals was 95.83% ( Figure 3 ).

[0075] 6. Blood Pressure Lowering Activity Detection

[0076] The angiotensin-converting enzyme 2 (ACE2) inhibitor screening kit of Beyotime was used to detect the antihypertensive activity of the hydrolyzed peptide.

[0077] The results showed that black chicken liver peptide has ACE2 inhibitory activity, with an inhibition rate of 12.35%. The specific detection steps are as follows:

[0078] 6.1 Preparation of test solution

[0079] Take 20mg of black-bone chicken liver hydrolyzed peptide powder and add 200μL Assay Buffer to prepare a 10mg / mL solution.

[0080] 6.2 Preparation of positive control

[0081] Take 1 μL of 0.5 mM positive control inhibitor mLN-4760 and add it to 999 μL Assay Buffer to prepare a 50 nM positive control inhibitor mLN-4760.

[0082] 6.3 Sample determination

[0083] 6.3.1 Preparation of Assay Reagent

[0084] Prepare an appropriate amount of Assay Reagent based on the number of samples (including relevant controls). Add 10 μL of ACE2 Enzyme to 1.85 mL of Assay Buffer and mix thoroughly.

[0085] 6.3.2 Sample addition

[0086] Use a 96-well blackboard to set up each group, and add the detection reagents and samples in sequence according to Table 6. After adding the sample to be tested, mix well. Set up 3 replicate wells for each sample. Quickly add 2μL of substrate to each well on ice and mix well. Incubate at 37℃ in the dark for 10 minutes and then use enzyme labeling for fluorescence measurement. The excitation wavelength is 325nm and the emission wavelength is 393nm. The average fluorescence value of each sample well and blank control well can be recorded as RFU respectively. 空白对照 、RFU 100％酶活性对照 、RFU 阳性对照 and RFU 样品The calculation formula of the inhibition percentage of the sample is as follows: Inhibition rate (%) = (RFU 100％酶活性对照 -RFU 样品 ) / (RFU 100％酶活性对照 -RFU 空白对照 )×100%.

[0087] Table 6 Sample addition method for antihypertensive activity detection

[0088]

[0089] 7. Animal Experimentation

[0090] 7.1 Grouping and Modeling

[0091] After one week of adaptive culture, all 5-week-old C57 mice were divided into five groups, with 10 mice in each group: normal group (NC): free access to drinking water and purified water; model group (M): 3.5% DSS and purified water; positive control group (ASA): 3.5% DSS and oral administration of 40 mg / kg 5-aminosalicylic acid; low-dose group (BLPL): 3.5% DSS and 100 mg / kg black-bone chicken liver hydrolyzed peptide; high-dose group (BLPH): 3.5% DSS and 300 mg / kg black-bone chicken liver hydrolyzed peptide. During the breeding period, mice were weighed daily, blood in stool was observed, and disease activity index (DAI) scores were recorded. After the end of the drug administration, mice were killed by cervical dislocation, and blood, colon and colon contents were collected, and colon length was measured.

[0092] 7.2 Mouse weight loss rate, colon length measurement and DAI score

[0093] During the experiment, mice were weighed daily before gavage. The average of the three weight loss scores was used to determine the final DAI score. The specific criteria are shown in Table 7. After sacrifice, the colon length of each mouse was measured. The weight loss rate was calculated using the following formula: Weight loss rate (%) = [mouse weight at that time (g) - mouse weight before modeling (g)] / mouse weight before modeling (g) × 100%.

[0094] Table 7 DAI scoring criteria

[0095]

[0096] Colon shortening is another significant feature of dextran sulfate sodium-induced colon inflammation. The NC and ASA groups had the longest colon length, approximately 8.0 cm; the DSS group had the shortest length, only about 4.5 cm. Compared with the DSS group, the colon length of the BLPL and BLPH groups increased to approximately 6.5 cm. It can be seen that there was no significant difference in the effect of different doses of black chicken liver peptide on colon length ( Figure 4 ).

[0097] The DAI score showed that there was no significant difference in scores between the groups in the first 4 days, but the score gap began to widen from the 5th day. The NC group always maintained a low level, the DSS group scored much higher than the other groups, and the ASA group, BLPL group and BLPH group had similar levels in the last 3 days, indicating that black chicken liver peptide did have an alleviating effect on colitis in mice ( Figure 5 ).

[0098] 7.3 Colonic Histopathological Observation

[0099] Colon tissue fixed in 4% paraformaldehyde solution was obtained, dehydrated, embedded, and sliced. The slices were then stained with HE and observed under an optical microscope to analyze the pathological damage.

[0100] Colon section results showed that the mucosal layer of the colon tissue in the DSS group was thinner, the crypt structure almost completely disappeared, and the inflammatory cell infiltration was severe; the mucosal layer of the BLPL and BLPH groups was thicker, the crypt structure was relatively intact, and the inflammatory cells were relatively few. Compared with the BLPL group, the colon tissue structure in the BLPH group was more intact and the inflammation level was lower. This shows that black chicken liver peptide can significantly reduce the colon tissue damage induced by dextran sulfate sodium in mice ( Figure 6 ).

[0101] 7.4 Determination of serum inflammatory factors

[0102] An appropriate amount of colon sample was taken, and pre-cooled physiological saline was added at a solid-liquid ratio of 1:9. After homogenization and centrifugation, the supernatant was collected. According to the instructions of the mouse interleukin IL-6, IL-1β, TNF-α and IL-10 ELISA quantitative detection kit (Shanghai ELISA Biotechnology Co., Ltd.), the contents of proinflammatory factors IL-6, IL-1β and TNF-α and the anti-inflammatory factor IL-10 were respectively determined.

[0103] Serum inflammatory factor detection showed that compared with the NC group, the levels of pro-inflammatory factors IL-6, IL-1β and TNF-α, and anti-inflammatory factor IL-10 in the DSS group were significantly increased, indicating that the inflammatory response in the DSS group was severe. Compared with the DSS group, the levels of inflammatory factors in the BLPL group and BLPH group decreased, indicating that black chicken liver peptide has an improving effect on DSS-induced enteritis ( Figure 7 ).

[0104] The results of the above experiments show that based on the colon length, DAI score and colon section results of mice, it can be concluded that the black chicken liver peptide prepared by the present invention has the effect of alleviating ulcerative colitis.

[0105] The embodiments described above are merely descriptions of preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Without departing from the spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by persons skilled in the art should fall within the scope of protection defined by the claims of the present invention.

Claims

1. An application of a black-bone chicken liver hydrolyzed peptide in the preparation of a drug having a blood pressure lowering effect, characterized in that: The preparation method of the black-bone chicken liver hydrolyzed peptide comprises the following steps: preparing the black-bone chicken liver into minced meat and mixing the mixture with water, heating the obtained liquid, adding alkaline protease for enzymatic hydrolysis, inactivating the enzyme, centrifuging and collecting the supernatant, and freeze-drying to obtain the black-bone chicken liver hydrolyzed peptide; preparing the black-bone chicken liver into minced meat and mixing the mixture with water such that the minced meat is adjusted to a mass concentration of 30% by adding water; adding the alkaline protease in an amount of 1% of the mass of the minced meat in the liquid; performing the enzymatic hydrolysis at 55°C for 1 hour; inactivating the enzyme at 90°C for 15 minutes; and centrifuging at 8000 r / min at 4°C for 10 minutes.

2. A use of a black-bone chicken liver hydrolyzed peptide in the preparation of a medicament for improving ulcerative colitis, characterized in that: The preparation method of the black-bone chicken liver hydrolyzed peptide comprises the following steps: preparing the black-bone chicken liver into minced meat and mixing the mixture with water, heating the obtained liquid, adding alkaline protease for enzymatic hydrolysis, inactivating the enzyme, centrifuging and collecting the supernatant, and freeze-drying to obtain the black-bone chicken liver hydrolyzed peptide; preparing the black-bone chicken liver into minced meat and mixing the mixture with water such that the minced meat is adjusted to a mass concentration of 30% by adding water; adding the alkaline protease in an amount of 1% of the mass of the minced meat in the liquid; performing the enzymatic hydrolysis at 55°C for 1 hour; inactivating the enzyme at 90°C for 15 minutes; and centrifuging at 8000 r / min at 4°C for 10 minutes.

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