A crocodile blood peptide complex for improving immunity and preparation method thereof

Abstract

The present invention provides a crocodile blood peptide complex for improving immunity and a preparation method thereof. The present invention obtains two polypeptides with good effects of promoting immune cell proliferation and improving immune activity from the blood of cultivated Siamese crocodiles through enzymatic hydrolysis, screening and identification. After preparing the polypeptides into a complex, the immune activity of mice can be well improved, and the polypeptides have good application value.

Description

Technical Field

[0001] The present application relates to the biological field, and to a crocodile blood peptide complex for improving immunity and a preparation method thereof. Background Art

[0002] The immune system is a vital part of the human body's ability to resist foreign pathogens and improve survival, but it is easily disturbed by many factors such as unhealthy lifestyles, germs, and stress. Once the human immune system has problems, immunosuppressants such as tacrolimus (FK506), cyclosporine, rapamycin, and mizoribine are usually used to regulate immune capacity. However, most immunosuppressants have slow effects and certain toxic side effects. In recent years, researchers have discovered that natural bioactive peptides may become a major breakthrough in the treatment of immune diseases.

[0003] The ocean contains huge biological resources, rich and diverse species and unique chemical substances. Marine organisms such as fish, shellfish, mollusks, crustaceans and their by-products are rich sources of high-quality protein. Peptides from marine organisms have antioxidant, blood pressure regulating, antibacterial, anti-cancer, and immune-enhancing functions. For example, two peptides with strong antioxidant activity were obtained from fermented fish sauce; the component less than 1kDa of bigeye tuna head protein has good scavenging ability for hydroxyl free radicals, superoxide anions, etc. and improves immunity.

[0004] In addition, plant protein rice protein mainly contains gluten, globulin and alcohol-soluble protein, and its main advantages are reasonable amino acid composition and low allergenicity. At present, rice protein peptides are mainly focused on the research of antioxidant and antihypertensive activities, but the research on rice immune active peptides has also received more and more attention. As early as 1994, Takahashi et al. isolated an octapeptide from rice pancreatic hydrolysate and proved that it has immunomodulatory effects such as enhancing macrophage phagocytosis, promoting intestinal contraction and anti-morphine. After that, some rice peptides with opioid activity were separated one after another. Recently, Wang Lu et al. found that rice peptides of different purities separated and purified by pancreatic hydrolysis can promote the proliferation of macrophages RAW164.7, and enzymatic peptides with a molecular weight of ≤1000Da have the best pro-proliferation effect. Studies have found that selenium-enriched rice protein hydrolysates can protect RAW264.7 macrophage toxicity caused by Pb, indicating that rice active peptides have an immunoprotective effect on immune cells.

[0005] Other plant-derived immunoactive peptides There are many types of food-derived plants, and fruits and vegetables have also become the main sources of immunoactive peptides. The immunological test of Actinidia arguta peptides showed that Actinidia arguta peptides can prolong the weight-bearing swimming time of mice, increase the liver glycogen reserve of mice, enhance the sports endurance of animals, and relieve the physical fatigue of animals, which has a direct immunoenhancing effect on the mouse body. The protein active peptide SSEDIKE isolated from Amaranth can inhibit the gene expression of chemokine CC120 in human colon cancer cells Caco-2, and has the immune characteristics of inhibiting inflammation. The cyclic peptide citrusinXl was isolated from mandarin oranges, and the RAW264.7 inflammation model was used to prove that it can reduce the production of iNOS and N by inhibiting the NF-xB pathway, and has an anti-inflammatory effect. In addition, the anti-inflammatory immunoactive peptide GPETAFLR from lupin can act on giant cells differentiated from the human monocytic cell line THP-1 and significantly reduce the pro-inflammatory mediators TNF, 1L-18 and CCL2, and increase the gene expression of the anti-inflammatory mediator IL-10. The research on these immune active peptides not only demonstrates the biological activity of food plants, but also provides more opportunities for the full utilization of food plants in production and the improvement of clinical diseases.

[0006] Fungal immunologically active peptides Fungal proteins are a special type of protein whose amino acid molecules are arranged linearly. Different proteins can be produced according to different bacterial species. Crude extracts of fungal proteins and peptides, polysaccharide peptides (PSP) and cyclic peptides isolated from them have immunomodulatory activity in vitro and in vivo. Antimicrobial peptides were isolated and extracted from Cordyceps sinensis and Pleurotus eryngii, respectively. These peptides have a significant inhibitory effect on the proliferation of various pathogenic bacteria. Studies have found that PSP of Coriolus versicolor can regulate the Toll-like receptor 4 and 5 signaling pathways of human peripheral blood monocytes, proving that Coriolus versicolor glycopeptides have in vitro immune activity. In vivo studies in rats, it was found that PSP of Coriolus versicolor can increase the levels of TNF-a and IL-6 in LPS-induced rat serum, proving that PSP has in vivo immunomodulatory function.

[0007] Crocodile peptides are rich in nutrients. They are high-protein, low-sugar, and low-fat products. They are rich in potassium, calcium, magnesium, zinc, iron and other metal elements. Moreover, they have a complete range of amino acids and are rich in 8 essential amino acids for the human body. In particular, the content of lysine and leucine, which are the most needed by the human body, is significantly higher. In addition, the ratio of branched-chain amino acids to aromatic amino acids in crocodile active peptides is significantly higher than that of various fish. Studies have shown that crocodile active peptides can effectively remove free radicals that are harmful to the human body, improve the body's antioxidant capacity, reduce the production of oxidation products, and protect the body from various oxidative damage, thereby preventing various diseases related to free radical damage, such as hypertension, diabetes, Alzheimer's disease, and cardiovascular disease. For example, crocodile skin collagen active peptides can effectively inhibit angiotensin-converting enzyme, thereby achieving the effect of lowering blood pressure; crocodile meat active peptides can repair damaged cells, improve pancreatic cell function, promote insulin secretion and the realization of pancreatic islet function, while improving the role of antioxidant enzymes, reducing oxidation, and reducing the occurrence of diabetic complications. Crocodile peptides are rich in antioxidants, which can neutralize free radicals, reduce cell damage, and thus enhance the function of the immune system. Antioxidants not only boost the vitality of immune cells, but also inhibit inflammatory responses, helping maintain overall health.

[0008] However, at present, on the one hand, there are not enough types of highly active crocodile peptides isolated, and on the other hand, the number of crocodiles is limited, the preparation method is complicated, the cost is high, and the source of peptides is limited. It is not suitable to directly obtain active peptides by isolating and preparing them from crocodile meat or crocodile blood. Summary of the invention

[0009] The invention provides a crocodile blood peptide complex for improving immunity.

[0010] Furthermore, the crocodile blood peptide complex of the present invention is composed of YZB-II-B-1 and YZB-II-B-2 polypeptides.

[0011] Furthermore, the amino acid sequence of the YZB-II-B-1 polypeptide is shown in SEQ ID NO: 1, and the amino acid sequence of the YZB-II-B-2 polypeptide is shown in SEQ ID NO: 2.

[0012] Further, the polypeptide of the present invention can also be substituted. Specifically, the amino acid substitution can be a conservative amino acid substitution. Conservative amino acid substitutions can be, for example, aspartic acid-glutamic acid as acidic amino acids; lysine / arginine / histidine as basic amino acids; leucine / isoleucine, methionine / valine, alanine / valine as hydrophobic amino acids; serine / glycine / alanine / threonine as hydrophilic amino acids. Conservative amino acid substitutions also include groupings based on side chains. For example, a group of amino acids with aliphatic side chains are glycine, alanine, valine, leucine and isoleucine; a group of amino acids with aliphatic-hydroxy side chains are serine and threonine; a group of amino acids with amide-containing side chains are asparagine and glutamine; a group of amino acids with aromatic side chains are phenylalanine, tyrosine and tryptophan; a group of amino acids with basic side chains are lysine, arginine and histidine; a group of amino acids with sulfur-containing side chains are cysteine ​​and methionine. After amino acid substitutions are made, the activity of the polypeptide containing the amino acid substitutions can be evaluated using the assays described herein.

[0013] Furthermore, the polypeptide of the present invention can also be coupled to a polymer to increase biological activity or half-life.

[0014] Specifically, the polymer molecule coupled to the polypeptide can be any suitable polymer molecule having a molecular weight as defined in the present invention, including natural and synthetic homopolymers, such as polyols (i.e., poly-OH), polyamines (i.e., poly-NH 2) and polycarboxylic acids (ie poly-COOH), and further, heteropolymers, ie polymers comprising one or more different coupling groups such as hydroxyl groups or amine groups. Examples of suitable polymer molecules include polymer molecules selected from the group consisting of polyalkylene oxides (PAOs), such as polyalkylene glycols (PAGs), including polyethylene glycol (PEG), methoxypolyethylene glycol (mPEG) and polypropylene glycol, PEG-glycidyl ether (Epox-PEG), PEG-oxycarbonyl imidazole (CDI-PEG) branched polyethylene glycols (PEGs), polyvinyl alcohol (PVA), polycarboxylates, polyvinyl pyrrolidone, poly-D,L-amino acids, polyethylene maleic anhydride copolymers, polystyrene malic anhydride copolymers, dextran including carboxymethyl-dextran, heparin, homologous albumin, cellulose including methylcellulose, carboxymethylcellulose, ethylcellulose, hydroxyethylcellulose, carboxyethylcellulose and hydroxypropylcellulose, hydrolysates of chitosan, starches such as hydroxyethyl-starch and hydroxypropyl-starch, glycogen, agarose and derivatives thereof, guar gum, pullulan, inulin, xanthan gum, carrageenan, pectin, alginic acid hydrolysate and biopolymers. Preferred polymer molecules are non-toxic polymer molecules, such as (m)polyethylene glycol (mPEG), which further require relatively simple chemistry to be covalently coupled to attachment groups on the enzyme surface.

[0015] Furthermore, the present invention also provides the use of the crocodile blood peptide complex composed of YZB-II-B-1 and YZB-II-B-2 polypeptides in the preparation of a drug for improving immunity.

[0016] Specifically, the medicine contains a suitable carrier or excipient or a suitable pharmaceutical excipient.

[0017] Further, the medicament may contain: diluents such as lactose, sucrose, calcium hydrogen phosphate or carboxymethylcellulose; lubricants such as magnesium stearate, calcium stearate and talc; and binders such as starch, natural gums, for example gum arabic, gelatin, glucose, molasses, polyvinyl pyrrolidine, cellulose and its derivatives, povidone, crospovidone and other such binders, as known to those skilled in the art. Liquid pharmaceutically administrable compositions can be prepared by, for example, dissolving, dispersing or mixing the active compound defined above and optional pharmaceutical adjuvants in a carrier, such as water, saline, aqueous dextrose, glycerol, ethylene glycol, ethanol and the like, to form a solution or suspension. If desired, the pharmaceutical composition to be administered may also contain a small amount of non-toxic auxiliary substances such as wetting agents, emulsifiers or solubilizers, pH buffers and the like, for example acetate, sodium citrate, cyclodextrin derivatives, sorbitan monolaurate, triethanolamine sodium acetate, triethanolamine oleate and other such agents. Methods for preparing such dosage forms are known or will be apparent to those skilled in the art. The composition or formulation to be administered contains an amount of active compound sufficient to improve the symptoms of the subject being treated. In addition, antibacterial or antifungal agents, including but not limited to thimerosal, may be present in the formulation. Another formulation provided herein is a 5-hydroxy-1-nitropropene succinate-containing formulation in the presence of EDTA, NaCl and CaCl 2 The present invention also provides a stabilized solution in which an effective amount of polypeptide is contained and lactose is added. 2 Another formulation provided herein, either lyophilized or as a stabilized solution, contains an effective amount of a polypeptide in a solution containing EDTA, NaCl and CaCl 2 middle.

[0018] Furthermore, the medicine can be an oral preparation or an injectable preparation.

[0019] Further, the preparation for parenteral administration can include common excipients, sterile water, saline, polyalkylene glycols such as polyethylene glycol, vegetable oils, hydrogenated naphthalene and combinations thereof. In some instances, biocompatible, biodegradable lactide polymers, lactide / glycolide copolymers, polyoxyethylene (polyoxethylene)-polyoxypropylene copolymers or combinations thereof can be used as excipients for controlled release of polypeptides in vivo. Other suitable parenteral administration systems that can be used include but are not limited to ethylene vinyl acetate copolymer particles, osmotic pumps, implantable infusion systems, liposomes and combinations thereof. The preparation for inhalation administration can include excipients such as lactose. The inhalation preparation can be an aqueous solution containing such as polyoxyethylene-9-lauryl ether, glycocholate, deoxycholate or combinations thereof, or they can be an oil solution used in the form of nasal drops. If desired, the composition containing the polypeptide provided herein can be formulated into a gel for intranasal application. The preparation for parenteral administration can include glycocholate for oral administration. For oral administration, tablets or capsules can be prepared using suitable methods and pharmaceutically acceptable excipients, such as binders (e.g., pregelatinized corn starch, polyvinyl pyrrolidone, or hydroxypropyl methylcellulose); fillers (e.g., lactose, microcrystalline cellulose, or calcium hydrogen phosphate); lubricants (e.g., magnesium stearate, talc, or silicon dioxide); disintegrants (e.g., potato starch or sodium starch glycolate); or wetting agents (e.g., sodium lauryl sulfate). Tablets can be coated using suitable methods. Orally administered preparations can be formulated to provide controlled release of the polypeptide.

[0020] Furthermore, the present invention also provides a health product for improving immunity, wherein the health product contains a crocodile blood peptide complex for improving immunity. The crocodile blood peptide complex of the present invention is composed of YZB-II-B-1 and YZB-II-B-2 polypeptides.

[0021] Furthermore, the present invention also provides the use of the crocodile blood peptide complex in the preparation of a health product for improving immunity. Specifically, the crocodile blood peptide complex is composed of YZB-II-B-1 and YZB-II-B-2 polypeptides. Beneficial Effects

[0022] The present invention provides a crocodile blood peptide complex for improving immunity and a preparation method thereof. The present invention obtains two polypeptides with good effects of promoting immune cell proliferation and improving immune activity from the blood of cultivated Siamese crocodiles through enzymatic hydrolysis, screening and identification. After preparing the polypeptides into a complex, the immune activity of mice can be well improved, and the polypeptides have good application value. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1Effects of each component on the proliferation of RAW264.7 cells DETAILED DESCRIPTION

[0024] Those skilled in the art can refer to the content of this article and appropriately improve the process parameters to achieve. It is particularly important to point out that all similar replacements and modifications are obvious to those skilled in the art, and they are all considered to be included in the present invention. The methods and applications of the present invention have been described by preferred embodiments, and relevant personnel can obviously change or appropriately change and combine the methods and applications described herein without departing from the content, spirit and scope of the present invention to achieve and apply the technology of the present invention. The methods, equipment, and materials in the following implementation cases, if not specifically stated, are all conventional methods, equipment, and materials in the art, and can be purchased from the market.

[0025] Example 1 Preparation of crocodile blood peptide

[0026] Take the blood of farmed Siamese crocodiles, add 4 times the amount of distilled water, adjust to the appropriate pH, and add the optimized enzymatic hydrolysis conditions: reaction temperature is 50℃, pH is 7.0, enzyme to substrate concentration ratio is 6%, substrate concentration is 7.5%, and reaction time is 85min. The product after the reaction is placed in a 95℃ water bath for 15min to inactivate the enzyme. The inactivated solution is centrifuged at 4℃ and 4500r / min for 30min. The supernatant is vacuum concentrated and freeze-dried to obtain the enzymatic hydrolysate. The Siamese crocodile blood enzymatic hydrolysate is desalted using D101 macroporous adsorption resin, and ultrafiltration is used with ultrafiltration membranes with molecular weight cutoffs of 3kDa and 1kDa to obtain three components: YZB-I (MW>3kDa), YZB-II (3kDa>MW>1kDa) and YZB-III (MW<1kDa).

[0027] RAW264.7 cell line (Cat. No.: HTX1760, Haodi Huatuo Biotechnology) was cultured in high-glucose DMEM medium containing 10% inactivated fetal bovine serum at 37°C and 5% CO 2 Culture in a constant temperature incubator and subculture every 2-3 days. Take cells in the logarithmic growth phase and adjust the cell concentration to 5×10 5 100 μL of each well was inoculated into a 96-well culture plate and incubated at 37°C and 5% CO 2 Culture in a constant temperature incubator for 2 hours. When the cells adhere to the wall, remove the supernatant, add 100 μL of three ultrafiltration fractions to each well to make the final concentration 1 μg / mL (prepared with high-glucose DMEM medium containing 10% fetal bovine serum), add culture medium without polypeptide components to the blank control group, and use 1 μg / mL LPS as the positive control group. Set up 3 replicates for each group. Incubate at 37°C and 5% CO 2After continuing to culture in a constant temperature incubator for 24 hours, add 10 μL of 5 mg / mL MTT working solution, continue to culture for 4 hours, discard the culture medium, carefully rinse with PBS 3 times, add 150 μL DMSO dissolving solution to each well, oscillate in the dark for 20 minutes to completely dissolve the formazan, use a microplate reader to measure the OD value at 570nm, and calculate the proliferation rate of RAW264.7 cells, proliferation rate (%) = (sample OD570nm / blank control group OD570nm) × 100.

[0028] from Figure 1 It can be seen that the YZB-II (3kDa>MW>1kDa) component has the strongest ability to promote cell proliferation. Compared with the blank control group, the proliferation rate reached (165.4±3.7)%. Therefore, this component was selected for further separation and purification.

[0029] Dissolve the ultrafiltered YZB-II in double distilled water to maintain a concentration of 1 mg / mL, add it to a pre-equilibrated Sephadex G-25 gel column (2.6× 80 cm), and elute it with distilled water at a rate of 1 mL / min. Collect one tube every 5 minutes, and detect the absorbance at 220 nm. Merge according to the peaks to obtain 5 components (YZB-II-A~E). The same RAW264.7 cell proliferation rate (%) detection method as mentioned above was used to detect the effects of the five components on RAW264.7 cell proliferation. The results showed that the YZB-II-B component showed the strongest proliferation effect.

[0030] The YZB-II-B component was subjected to RP-HPLC. The RP-HPLC conditions were: chromatographic column Zorbax SB C-18 (4.6×250mm, 5μm, Agilent, USA); flow rate 1mL / min; detection wavelength 220nm; mobile phase: acetonitrile / water gradient elution; column temperature 25℃. YZB-II-B was analyzed by RP-HPLC as two peaks, named YZB-II-B-1 and YZB-II-B-2 respectively. The sequence of YZB-II-B-1 was determined by protein sequence analyzer using Edman degradation method as shown in SEQ ID NO: 1, and the sequence of YZB-II-B-2 was shown in SEQ ID NO: 2.

[0031] Example 2 Effect of crocodile blood peptide on phagocytic activity of immune cells

[0032] RAW264.7 cell line (Cat. No.: HTX1760, Haodi Huatuo Biotechnology) was cultured in high-glucose DMEM medium containing 10% inactivated fetal bovine serum at 37°C and 5% CO 2 Culture in a constant temperature incubator and subculture every 2-3 days. Take cells in the logarithmic growth phase and adjust the cell concentration to 5×105 A blank control group (medium only), a positive control group (1 μg / mL LPS), and two experimental groups (10 μg / mL and 100 μg / mL) of YZB-II-B-1 and YZB-II-B-2 were set up, with 4 replicate wells in each group. The cells were incubated at 37°C and 5% CO 2 After culturing for 24 h under the same conditions, the supernatant was discarded, 100 uL of 0.075% neutral red solution was added, and the cells were incubated at 37°C and 5% CO 2 Continue to culture in the incubator for 1 hour, discard the neutral red solution, carefully wash twice with PBS, add 100 μL / well of acetic acid-ethanol (1:1, V / V) cell lysis solution, let stand overnight at room temperature, use a microplate reader to measure the OD value at a wavelength of 540 nm, and calculate the phagocytic index of RAW264.7 cells according to the following formula. Phagocytic index (%) = (sample OD540nm / blank control group OD540nm) × 100. The results are shown in Table 1.

[0033] Table 1 Phagocytic index results of each group

[0034]

[0035] # indicates significant difference compared with the blank control group (P<0.05).

[0036] As can be seen from Table 1, after stimulating RAW264.7 cells, the YZB-II-B-1 and YZB-II-B-2 polypeptides of the present invention have a significant promoting effect on the phagocytic ability of cells, and have a certain dose dependence. Compared with the blank control group, the difference is significant (P<0.05).

[0037] Example 3 Effect of crocodile blood polypeptide on immunity of mice

[0038] BALB / c mice were randomly divided into 6 groups with 10 mice in each group: control group, model group, positive control group (fish scale collagen peptide 400 mg / kg) group (Cat. No. XCSW2022, Vanilla Bio), YZB-II-B-1 and YZB-II-B-2 (1:1) peptides totaling 200, 400, and 800 mg / kg groups. At the same time as modeling, oral administration began, once a day, for 15 consecutive days. The control group was given drinking water 0.2 mL / 10 g body weight. Modeling method Except for the control group, the mice in the other groups were intraperitoneally injected with cyclophosphamide, 0.6 mg / time, once a day, for a total of 3 times, and the control group was intraperitoneally injected with an equal amount of saline 0.2 mL / 10 g body weight. Mice were gavaged for 15 consecutive days, and then killed by cervical dislocation. The spleen was obtained under sterile conditions. The spleen was gently torn into pieces with forceps in a small dish containing an appropriate amount of sterile Hanks solution, and filtered through a 200-mesh sieve to prepare a single-cell suspension. The suspension was divided into two parts for use in transformation experiments. The cell suspension was washed twice with Hanks solution, centrifuged for 10 minutes (1000 r / min) each time, and then the cells were suspended in 1 mL of complete culture medium. The number of live cells was counted by trypan blue staining (above 95%), and the cell concentration was adjusted to 3×10 6 / mL. Each cell suspension was added to two wells of a 24-well culture plate, 1mL per well, 75μL ConA solution was added to one well as an experiment, and the other well was used as a control, and cultured in an incubator for 72h. 4h before the end of the culture, 0.7mL of supernatant was aspirated from each well, 0.7mL of serum-free RPMI1640 culture medium was added, and MTT 50μL / well was added, and the culture was continued for 4h. After the culture was completed, 1mL of acidic isopropanol was added to each well, and the mixture was blown to completely dissolve the purple crystals. Then it was dispensed into a 96-well culture plate, 3 parallel wells were made in each well, and the optical density value was measured with an enzyme reader at a wavelength of 570nm. The difference between the OD of the experimental well and the control well represents the proliferation capacity of lymphocytes. The results are shown in Table 2.

[0039] Table 2 Effects of each group on the proliferation of spleen lymphocytes in mice with immune impairment

[0040]

[0041] *Indicates P < 0.05 compared with the model group.

[0042] The results in Table 2 show that the proliferation function of T lymphocytes in the drug-treated mice was significantly higher than that in the model group, and was dose-dependent. Under the condition of 800 mg / kg administration, the proliferation ability was the strongest.

[0043] Furthermore, the spleen cell suspension (5×10 6 / L) was added into a 96-well cell culture plate, 100 μL / well. YAC-1 target cells (51 Cr-labeled target cells) were centrifuged and the cell density was adjusted to an effector-target ratio of 10:1. The cells were added to a 96-well culture plate and placed at 37°C and CO 2 The culture plate was incubated for 4 hours. The culture plate was centrifuged at 1000 rpm for 10 minutes, 100 μl of the supernatant was aspirated into a test tube, and the radioactivity per minute (cpm value) was measured using a liquid scintillation counter to calculate the killing rate of NK cells against target cells. The results are shown in Table 3.

[0044] Table 3 Effects of each group on NK cell killing activity in mice with immune damage

[0045]

[0046] *Indicates P < 0.05 compared with the model group.

[0047] The NK cell killing results of different groups in Table 3 show that the positive control group and the polypeptide group of the present invention have positive immunomodulatory effects, can improve the immune function of mice, and are dose-dependent. Based on the above results, it can be found that the composition drug prepared by the YZB-II-B-1 and YZB-II-B-2 polypeptides of the present invention can effectively improve the body's immunity and has excellent therapeutic effects and application prospects.

[0048] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein by equivalents. However, these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A crocodile polypeptide for improving immunity, characterized in that It is named as YZB-II-B-1 polypeptide, and its amino acid sequence is shown in SEQ ID NO:

1.

2. A crocodile polypeptide for improving immunity, characterized in that It is named as YZB-II-B-2 polypeptide, and its amino acid sequence is shown in SEQ ID NO:

2.

3. A crocodile blood peptide complex for improving immunity, characterized in that The complex is composed of YZB-II-B-1 and YZB-II-B-2 polypeptides in a 1:1 ratio, wherein the amino acid sequence of the YZB-II-B-1 polypeptide is shown in SEQ ID NO: 1, and the amino acid sequence of the YZB-II-B-2 polypeptide is shown in SEQ ID NO:

2.

4. Use of crocodile blood peptide complex in the preparation of drugs for improving immunity, wherein: The complex is composed of YZB-II-B-1 and YZB-II-B-2 polypeptides in a 1:1 ratio, wherein the amino acid sequence of the YZB-II-B-1 polypeptide is shown in SEQ ID NO: 1, and the amino acid sequence of the YZB-II-B-2 polypeptide is shown in SEQ ID NO: 2.

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