Calf thymosin capable of improving immunity as well as preparation method and application of calf thymosin

Abstract

The invention discloses calf thymosin for improving immunity as well as a preparation method and application thereof, and relates to the technical field of functional foods. The preparation method comprises the following steps: mixing a calf thymus raw material with an auxiliary agent and water, adjusting the pH value to 4.5-6, and carrying out ball milling treatment to obtain a raw material 1; mixing the raw material 1 with water, cooking and filtering to obtain a raw material 2; mixing the raw material 2 with a first compound enzyme for enzymolysis to obtain an enzymatic hydrolysate 1; the enzymatic hydrolysate 1 and a second compound enzyme are mixed for enzymolysis to obtain enzymatic hydrolysate 2, and the enzymatic hydrolysate 2 is subjected to ultrafiltration concentration and drying to obtain the calf thymosin. By optimizing the extraction process of calf thymosin, the calf thymosin product with high content, high activity and better quality is obtained.

Description

Technical Field

[0001] The invention relates to the technical field of functional foods, and in particular to a calf thymosin peptide for improving immunity, and a preparation method and application thereof. Background Art

[0002] The body's immunity can also be called resistance in general, which refers to the body's ability to resist external invasion, such as bacteria and viruses. When the body is tired or the body's functions are not coordinated, the body's immunity will decline.

[0003] Clinically, the occurrence of diseases such as influenza, rhinitis, cough, allergic dermatitis, asthma, and peptic ulcer are all related to decreased immunity. Since the early symptoms of decreased immunity are not obvious and difficult to detect, once the body shows obvious diseases, it means that the immunity has been greatly reduced, which will bring great difficulties to the treatment of the disease and the patient's recovery. Aging is an important reason for the decline of human immunity. In addition, in addition to the decline of immunity caused by aging, bad living habits can also harm people's immunity. For example, staying up late, smoking, alcoholism, lack of exercise, etc., will lead to decreased immunity. Therefore, no matter what age group people are at, improving immunity is the key to staying healthy.

[0004] The thymus gland is one of the important immune organs of the body. Thymosin is a group of polypeptide hormones secreted by the thymus gland, which mainly include two types of proteins with molecular weights of about 9600 and 7000. It has therapeutic effects on a variety of body diseases, such as various types of severe hepatitis, myasthenia gravis, systemic lupus erythematosus, rheumatoid arthritis, viral pneumonia, and it also has a good auxiliary therapeutic effect on tumors. It is currently mainly used in products that improve immunity. For example, Chinese patent CN115299545A discloses a solid beverage for improving human immunity, comprising the following components by weight: 80-120 parts of composite polypeptide, 10-25 parts of tuckahoe polysaccharide, 10-25 parts of yeast β-glucan, and 30-40 parts of bovine colostrum powder; the composite polypeptide comprises one or more of tuckahoe peptide powder, bovine collagen peptide, ginseng peptide powder, and calf thymus peptide powder; and the preparation method thereof is: after the composite polypeptide, tuckahoe polysaccharide, yeast β-glucan, and bovine colostrum powder are mixed, dried, and sterilized, a solid beverage for improving human immunity is obtained. The solid beverage for improving human immunity of the application has the advantages of promoting human metabolism, enhancing the efficacy of protein, promoting material absorption, and improving human immunity and anti-fatigue ability.

[0005] At present, the calf thymosin peptide preparations produced domestically are all made from calf thymus as raw material to extract highly active thymosin peptides. The extraction processes include acid extraction method, neutral extraction method, neutral homogenate freeze-thaw extraction method, etc.

[0006] Chinese patent CN 112450316 A discloses a method for preparing calf thymus peptide powder, which includes grinding 4 times → homogenizing 5 times → freeze-thawing 3 times → sterilization → enzymolysis → filtration → concentration → filtration → spray drying. Through grinding, homogenization, freeze-thawing and enzymolysis, small molecular peptides with small molecular weight and high peptide content are fully extracted and retained, and high-quality calf thymus peptide powder with small molecular weight, high clarity and good taste is obtained through centrifugation and ceramic membrane filtration. After testing, the peptide content of the final product is as high as 98.7%, and the molecular weight distribution below 1000 Dalton accounts for 73.7%.

[0007] Currently, there is a great demand for functional foods that can enhance immunity, and there is an urgent need for a method for extracting calf thymosin with high content, high activity and stability. Summary of the invention

[0008] In view of this, the purpose of the present invention is to provide a calf thymosin peptide for improving immunity and a preparation method and application thereof, and to obtain a calf thymosin peptide product with high content, high activity and better quality by optimizing the extraction process of calf thymosin peptide.

[0009] To achieve the above-mentioned object of the invention, the technical solution of the present invention is as follows:

[0010] In one aspect, the present invention provides a method for preparing calf thymosin for improving immunity, comprising the steps of:

[0011] (1) ball milling: calf thymus raw material is mixed with an auxiliary agent and water, the pH is adjusted to 4.5-6, and ball milling is performed to obtain raw material 1; the auxiliary agent is a mixture of calcium chloride and colloidal silicon dioxide;

[0012] (2) Steaming: mixing the raw material 1 obtained in step (1) with water, steaming, and filtering to obtain raw material 2;

[0013] (3) the first step of enzymatic hydrolysis: the raw material 2 obtained in step (2) is mixed with a first composite enzyme for enzymatic hydrolysis to obtain an enzymatic hydrolyzate 1; the first composite enzyme is a mixture of lipase and alkaline protease;

[0014] (4) second enzymatic hydrolysis: the enzymatic hydrolyzate 1 obtained in step (3) is mixed with a second composite enzyme for enzymatic hydrolysis to obtain an enzymatic hydrolyzate 2; the second composite enzyme is a mixture of papain, bromelain and carboxypeptidase;

[0015] (5) Concentration: The enzymatic hydrolysate 2 is concentrated by ultrafiltration and dried to obtain the calf thymosin peptide.

[0016] Preferably, in step (1), the mass ratio of calcium chloride to colloidal silicon dioxide is 1-2:1-2, more preferably 1:1.

[0017] More preferably, the particle size of the colloidal silica is less than 200 mesh.

[0018] Preferably, in step (1), the mass ratio of the calf thymus raw material, the auxiliary agent and water is 5-15:1-5:15-25, and more preferably 10:2:20.

[0019] Preferably, in step (1), the pH is adjusted to 5-5.5, more preferably 5.

[0020] Preferably, in step (1), the ball milling conditions are: filling rate 70-90%, rotation speed 1500-3000 r / min, and ball milling time 30-60 min; more preferably: filling rate 80%, rotation speed 2000 r / min, and ball milling time 40 min.

[0021] Preferably, in step (2), the cooking conditions are: the pressure is maintained at 0.25±2Mpa, the temperature is maintained at 120±5°C, and the cooking is carried out for 3-8h; more preferably, the pressure is maintained at 0.25±2Mpa, the temperature is maintained at 120±5°C, and the cooking is carried out for 4h.

[0022] Preferably, in step (3), the mass ratio of the lipase to the alkaline protease is 1:2-5, more preferably 1:3.

[0023] Preferably, in step (3), the conditions for the first enzymatic hydrolysis are: enzymatic hydrolysis temperature is 45-55°C, and the time is 0.5-1.5h; more preferably: enzymatic hydrolysis temperature is 50°C, and the time is 1h.

[0024] Preferably, in step (3), the mass of the first complex enzyme added is 1-3% of the mass of raw material 2, and more preferably 2%.

[0025] Preferably, in step (4), the mass ratio of papain, bromelain and carboxypeptidase is 1:0.5-2:1-5, more preferably 1:1:2.

[0026] Preferably, in step (4), the conditions for the second step of enzymatic hydrolysis are: enzymatic hydrolysis temperature is 40-50°C, and the time is 30-60 min; more preferably: enzymatic hydrolysis temperature is 45°C, and the time is 40 min.

[0027] Preferably, in step (4), the mass of the second complex enzyme added is 0.5-2% of the mass of raw material 2, and more preferably 1%.

[0028] Preferably, in step (5), in the ultrafiltration concentration, the molecular weight of the ultrafiltration membrane is 10,000.

[0029] As a specific example of the present invention, the preparation method comprises the steps of:

[0030] (1) Pretreatment: Wash and mince the calf thymus raw material, mix it with an auxiliary agent (a mixture of calcium chloride and colloidal silicon dioxide (particle size <200 mesh) in a mass ratio of 1:1) and water, the mass ratio of the calf thymus raw material, auxiliary agent and water being 10:2:20, adjust the pH to 5, add it into a ball mill for ball milling, the filling rate is 80%, the rotation speed is 2000 r / min, and the ball milling is carried out for 40 minutes to obtain raw material 1;

[0031] (2) Mix the raw material 1 obtained in step (1) with 3 times the weight of water, add it into a cooking tank, maintain the pressure at 0.25±2Mpa, maintain the temperature at 120±5°C, cook for 4 hours, filter, collect the cooking liquid, cool it, and obtain raw material 2;

[0032] (3) Adding the first complex enzyme to the raw material 2 obtained in step (2), the mass of the first complex enzyme added is 2% of the raw material 2, adjusting the pH to 8.0, mixing and enzymolysis, and obtaining an enzymolysis solution 1; wherein the first complex enzyme is a mixture of lipase and alkaline protease in a mass ratio of 1:3, the enzymolysis temperature is 50°C, and the time is 1 hour.

[0033] (4) Second step of enzymolysis: the pH of the enzymolysis solution 1 obtained in step (3) is adjusted to 7.0, and a second composite enzyme is added for mixed enzymolysis, wherein the mass of the second composite enzyme added is 1% of the raw material 2, to obtain an enzymolysis solution 2; wherein the second composite enzyme is a mixture of papain, bromelain and carboxypeptidase in a mass ratio of 1:1:2, the enzymolysis temperature is 45°C, and the time is 40 min.

[0034] (5) Concentration: The enzymatic hydrolysate 2 is concentrated by ultrafiltration (ultrafiltration membrane molecular weight 10000), sterilized, and spray-dried to obtain the calf thymosin peptide.

[0035] On the other hand, the present invention provides calf thymosin peptide prepared by the above preparation method.

[0036] Finally, the present invention provides the use of the calf thymosin peptide prepared by the above preparation method in the preparation of products for improving immunity.

[0037] The beneficial effects of the present invention are:

[0038] (1) The present invention optimizes the extraction process of calf thymosin and obtains a calf thymosin product with high content, high activity and greater stability through ball milling and two-step enzymatic hydrolysis.

[0039] (2) The present invention selects specific ball milling aids, conditions and types of complex enzymes for the extraction of calf thymosin, which significantly improves the quality of the product. DETAILED DESCRIPTION

[0040] In order to make the technical means, creative features, purpose and efficacy of the present invention easy to understand, the present invention is further explained in conjunction with specific embodiments below, but the following embodiments are only preferred embodiments of the present invention, not all. Based on the embodiments in the implementation mode, other embodiments obtained by those skilled in the art without creative work all belong to the protection scope of the present invention. In the following embodiments, unless otherwise specified, the operating methods used are conventional operating methods, the equipment used are conventional equipment, and the equipment materials used in each embodiment are the same.

[0041] In the following embodiments, the lipase activity is 100,000 U / g, the papain activity is 100,000 U / g, the alkaline protease activity is 100,000 U / g, the papain activity is 100,000 U / g, the bromelain activity is 100,000 U / g, the carboxypeptidase activity is 200,000 U / g, the neutral protease activity is 200,000 U / g, and the colloidal silica is Hepu Youxin Colloidal Silica 200PHARM.

[0042] Example 1

[0043] (1) Pretreatment: The calf thymus raw material was washed and minced, and mixed with an auxiliary agent (a mixture of calcium chloride and colloidal silicon dioxide (particle size <200 mesh) in a mass ratio of 1:1) and water. The mass ratio of the calf thymus raw material, auxiliary agent and water was 10:2:20. The pH was adjusted to 5.0 with citric acid, and the raw material was added to a ball mill for ball milling. The ball milling conditions were: filling rate 80%, rotation speed 2000 r / min, and ball milling for 40 min, to obtain raw material 1;

[0044] (2) Mix the raw material 1 obtained in step (1) with 3 times its weight of water, add it into a cooking tank, maintain the pressure at 0.25±2Mpa, maintain the temperature at 120±5°C, cook for 4 hours, filter, collect the cooking liquid, cool it, and obtain raw material 2;

[0045] (3) Adding the first complex enzyme to the raw material 2 obtained in step (2), the mass of the first complex enzyme added is 2% of the raw material 2, adjusting the pH to 8.0, mixing and enzymolysis, and obtaining an enzymolysis solution 1; wherein the first complex enzyme is a mixture of lipase and alkaline protease in a mass ratio of 1:3, the enzymolysis temperature is 50°C, and the time is 1 hour.

[0046] (4) Second step of enzymolysis: the enzymatic solution 1 obtained in step (3) is inactivated by high temperature, the pH value is adjusted to 7.0 after cooling, and the second composite enzyme is added for mixed enzymolysis, wherein the mass of the second composite enzyme added is 1% of the raw material 2, to obtain an enzymatic solution 2; wherein the second composite enzyme is a mixture of papain, bromelain and carboxypeptidase in a mass ratio of 1:1:2, the enzymatic hydrolysis temperature is 45°C, and the time is 40 min.

[0047] (5) Concentration: The enzymatic hydrolysate 2 is concentrated by ultrafiltration (ultrafiltration membrane molecular weight 10000), sterilized, and spray-dried to obtain the calf thymosin peptide.

[0048] Example 2

[0049] (1) Pretreatment: The calf thymus raw material was washed and minced, and mixed with an auxiliary agent (a mixture of calcium chloride and colloidal silicon dioxide (particle size <200 mesh) in a mass ratio of 2:1) and water. The mass ratio of the calf thymus raw material, auxiliary agent and water was 10:1:20. The pH was adjusted to 4.2 with citric acid, and the raw material was added to a ball mill for ball milling. The ball milling conditions were: filling rate 80%, rotation speed 2000 r / min, and ball milling for 40 min, to obtain raw material 1;

[0050] (2) Mix the raw material 1 obtained in step (1) with 3 times its weight of water, add it into a cooking tank, maintain the pressure at 0.25±2Mpa, maintain the temperature at 120±5°C, cook for 4 hours, filter, collect the cooking liquid, cool it, and obtain raw material 2;

[0051] (3) Adding the first complex enzyme to the raw material 2 obtained in step (2), the mass of the first complex enzyme added is 1% of the raw material 2, adjusting the pH to 8.0, mixing and enzymolysis to obtain an enzymolysis solution 1; wherein the first complex enzyme is a mixture of lipase and alkaline protease in a mass ratio of 1:2, the enzymolysis temperature is 50°C, and the time is 1 hour.

[0052] (4) Second step of enzymolysis: the enzymatic solution 1 obtained in step (3) is inactivated by high temperature, the pH value is adjusted to 7.0 after cooling, and the second composite enzyme is added for mixed enzymolysis, wherein the mass of the second composite enzyme added is 2% of the raw material 2, to obtain an enzymatic solution 2; wherein the second composite enzyme is a mixture of papain, bromelain and carboxypeptidase in a mass ratio of 1:2:5, the enzymatic hydrolysis temperature is 45°C, and the time is 40 min.

[0053] (5) Concentration: The enzymatic hydrolysate 2 is concentrated by ultrafiltration (ultrafiltration membrane molecular weight 10000), sterilized, and spray-dried to obtain the calf thymosin peptide.

[0054] Example 3

[0055] (1) Pretreatment: The calf thymus raw material was washed and minced, and mixed with an auxiliary agent (a mixture of calcium chloride and colloidal silicon dioxide (particle size <200 mesh) in a mass ratio of 1:2) and water. The mass ratio of the calf thymus raw material, auxiliary agent and water was 10:5:20. The pH was adjusted to 6.0 with citric acid, and the raw material was added to a ball mill for ball milling. The ball milling conditions were: filling rate 80%, rotation speed 2000 r / min, and ball milling for 40 min, to obtain raw material 1;

[0056] (2) Mix the raw material 1 obtained in step (1) with 3 times its weight of water, add it into a cooking tank, maintain the pressure at 0.25±2Mpa, maintain the temperature at 120±5°C, cook for 4 hours, filter, collect the cooking liquid, cool it, and obtain raw material 2;

[0057] (3) adding a first complex enzyme to the raw material 2 obtained in step (2), wherein the mass of the first complex enzyme added is 3% of the raw material 2, adjusting the pH to 8.0, mixing and enzymolysis to obtain an enzymolysis solution 1; wherein the first complex enzyme is a mixture of lipase and alkaline protease in a mass ratio of 1:2, the enzymolysis temperature is 50°C, and the time is 1 hour.

[0058] (4) Second step of enzymolysis: the enzymolysis solution 1 obtained in step (3) is inactivated by high temperature, the pH value is adjusted to 7.0 after cooling, and the second composite enzyme is added for mixed enzymolysis, wherein the mass of the second composite enzyme added is 0.5% of the raw material 2 to obtain an enzymolysis solution 2; wherein the second composite enzyme is a mixture of papain, bromelain and carboxypeptidase in a mass ratio of 1:0.5:1, the enzymolysis temperature is 45°C, and the time is 40 min.

[0059] (5) Concentration: The enzymatic hydrolysate 2 is concentrated by ultrafiltration (ultrafiltration membrane molecular weight 10000), sterilized, and spray-dried to obtain the calf thymosin peptide.

[0060] Example 4

[0061] The difference from Example 1 is that the ball milling time is 30 min, and the rest are the same.

[0062] Example 5

[0063] The difference from Example 1 is that the ball milling time is 60 min, and the rest are the same.

[0064] Comparative Example 1

[0065] The difference from Example 1 is that the ball milling time is 20 min, and the rest are the same.

[0066] Comparative Example 2

[0067] The difference from Example 1 is that the ball milling time is 80 min, and the rest are the same.

[0068] Comparative Example 3

[0069] (1) Pretreatment: Wash and mince the calf thymus raw material to obtain raw material 1;

[0070] (2) Mix the raw material 1 obtained in step (1) with 3 times its weight of water, add it into a cooking tank, maintain the pressure at 0.25±2Mpa, maintain the temperature at 120±5°C, cook for 4 hours, filter, collect the cooking liquid, cool it, and obtain raw material 2;

[0071] (3) Adding the first complex enzyme to the raw material 2 obtained in step (2), the mass of the first complex enzyme added is 2% of the raw material 2, adjusting the pH to 8.0, mixing and enzymolysis, and obtaining an enzymolysis solution 1; wherein the first complex enzyme is a mixture of lipase and alkaline protease in a mass ratio of 1:3, the enzymolysis temperature is 50°C, and the time is 1 hour.

[0072] (4) Second step of enzymolysis: the enzymatic solution 1 obtained in step (3) is inactivated by high temperature, the pH value is adjusted to 7.0 after cooling, and the second composite enzyme is added for mixed enzymolysis, wherein the mass of the second composite enzyme added is 1% of the raw material 2, to obtain an enzymatic solution 2; wherein the second composite enzyme is a mixture of papain, bromelain and carboxypeptidase in a mass ratio of 1:1:2, the enzymatic hydrolysis temperature is 45°C, and the time is 40 min.

[0073] (5) Concentration: The enzymatic hydrolysate 2 is concentrated by ultrafiltration (ultrafiltration membrane molecular weight 10000), sterilized, and spray-dried to obtain the calf thymosin peptide.

[0074] Comparative Example 4

[0075] The difference from Example 1 is that the ball milling is changed to a split colloid mill for grinding for 40 min at a rotation speed of 2000 r / min, and the rest are the same.

[0076] Comparative Example 5

[0077] The difference from Example 1 is that in step (1), the pH is not adjusted, and the rest is the same.

[0078] Comparative Example 6

[0079] The difference from Example 1 is that no additive is added during ball milling, and the rest is the same.

[0080] Comparative Example 7

[0081] The difference from Example 1 is that colloidal silica is not added during ball milling, and the rest is the same.

[0082] Comparative Example 8

[0083] The difference from Example 1 is that in step (4), the second complex enzyme is a composition of papain, bromelain and neutral protease in a mass ratio of 1:1:2, and the rest are the same.

[0084] Comparative Example 9

[0085] The difference from Example 1 is that in step (3), the mass ratio of lipase to alkaline protease is 1:1, and the rest are the same.

[0086] Results

[0087] 1. Molecular weight distribution detection

[0088] According to the AGPC / UV test in Appendix 1 of GB 31645-2018, the proportion of peptides with a relative molecular weight not exceeding 2000 was counted. The calculation results of the examples and comparative examples are as follows:

[0089]

[0090] 2. Protein and peptide content detection

[0091] The protein content was determined according to the first method of GB5009.5-2016; the peptide content was determined according to Appendix BB of GB22492-2008.

[0092] The results are as follows:

[0093]

[0094]

[0095] 3. Biological activity detection

[0096] The activity was tested using the T lymphocyte-E rosette test (reference: Cai Hongjie, "Improvement of calf thymosin production process and analysis of preparations", No. 1672-0369 (2011) 19-2414-04), and the results are as follows:

[0097] Group Ea-RFC(%) Comparison 35 Example 1 73 Example 2 70 Example 3 72 Example 4 74 Example 5 72 Comparative Example 1 68 Comparative Example 2 65 Comparative Example 3 64 Comparative Example 4 66 Comparative Example 5 71 Comparative Example 6 68 Comparative Example 7 64 Comparative Example 8 67 Comparative Example 9 68

[0098] The test results of molecular weight distribution detection, protein and peptide content detection, and biological activity detection show that the calf thymosin peptide prepared in Examples 1-5 of the present invention has an increased content of short peptide chains, a higher peptide content, and a higher activity. Example 1, Example 4, Example 5, Comparative Example 1, and Comparative Example 2 screened the effect of ball milling time on product performance. The results showed that when the ball milling time was 30-60min, the calf thymosin peptide had better comprehensive performance. If the ball milling time was too short, the short peptide chain content and the peptide content would be reduced. If the ball milling time was too long, the activity would be significantly reduced. The results of Comparative Examples 3 and 4 show that ball milling pretreatment can greatly promote the subsequent enzymatic hydrolysis process and obtain products with smaller molecular weight and higher peptide content. Comparative Examples 5-7 screened the ball milling environment parameters and found that under certain acidic conditions and the action of composite additives, the comprehensive performance of calf thymosin peptide can be significantly improved. The results of Comparative Examples 8-9 show that a specific proportion of composite enzymes has a key influence on the quality of calf thymosin peptide.

[0099] 4. Improve immunity

[0100] (1) Experimental drugs

[0101] Calf thymosin peptide prepared in Example 1 and Comparative Example 3;

[0102] Spleen aminopeptide (National Medicine Standard No. H10970214).

[0103] (2) Experimental methods

[0104] Male Kunming mice (18-22g) were selected for adaptive feeding for one week, and the feeding environment temperature was 20±2℃, and the relative humidity was 45%-65%. They were randomly divided into groups, with 10 mice in each group. The experimental groups were respectively administered calf thymosin 500mg / kg / d of Example 1, calf thymosin 500mg / kg / d of Comparative Example 3, and spleen aminopeptide 1.3g / kg / d, and were intragastrically administered once a day (gastric volume 10mL / kg). The model group and the blank group were intragastrically administered with an equal volume of normal saline. After continuous administration for 2 weeks, each experimental group and the model group were intraperitoneally injected with cyclophosphamide 40mg / kg for two consecutive days for immunosuppression. After continuing to intragastrically administer for 2 weeks, the mice were killed by dislocation, the thymus and spleen were removed, and the thymus index and spleen index were calculated. The results are as follows:

[0105] Group Thymus index Spleen index Blank Group 0.244±0.125 0.310±0.354 Model Group 0.127±0.152** 0.224±0.266* Spleen aminopeptide group 0.263±0.201## 0.382±0.448## Example 1 0.258±0.247## 0.369±0.496## Comparative Example 3 0.218±0.138# 0.315±0.324#

[0106] In the table, compared with the blank group, the model group *P<0.05, **P<0.01; compared with the model group, the experimental group #P<0.05, ##P<0.01.

[0107] The results show that the calf thymosin peptide prepared in Example 1 of the present invention can significantly improve the thymus index and spleen index of immunosuppressive model mice, and the results are statistically significant. The calf thymosin peptide prepared in Comparative Example 3 has a significant effect of improving immunity, but there is still a gap compared with Example 1 and the spleen aminopeptide group. After process improvement, the present invention further improves the effect of calf thymosin peptide in improving immunity, and can be used to prepare products for improving immunity.

[0108] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for preparing calf thymosin peptide for improving immunity, characterized in that: Includes steps: (1) ball milling: calf thymus raw material is mixed with an auxiliary agent and water, the pH is adjusted to 4.5-6, and ball milling is performed to obtain raw material 1; the auxiliary agent is a mixture of calcium chloride and colloidal silicon dioxide; (2) Steaming: mixing the raw material 1 obtained in step (1) with water, steaming, and filtering to obtain raw material 2; (3) the first step of enzymatic hydrolysis: the raw material 2 obtained in step (2) is mixed with a first composite enzyme for enzymatic hydrolysis to obtain an enzymatic hydrolyzate 1; the first composite enzyme is a mixture of lipase and alkaline protease; (4) second enzymatic hydrolysis: the enzymatic hydrolyzate 1 obtained in step (3) is mixed with a second composite enzyme for enzymatic hydrolysis to obtain an enzymatic hydrolyzate 2; the second composite enzyme is a mixture of papain, bromelain and carboxypeptidase; (5) Concentration: The enzymatic hydrolysate 2 is concentrated by ultrafiltration and dried to obtain the calf thymosin peptide.

2. The preparation method according to claim 1, characterized in that: In step (1), the mass ratio of calcium chloride to colloidal silicon dioxide is 1-2:1-2.

3. The preparation method according to claim 1, characterized in that: In step (1), the mass ratio of the calf thymus raw material, the auxiliary agent and water is 5-15:1-5:15-25.

4. The preparation method according to claim 1, characterized in that: In step (1), the pH is adjusted to 5.

5. The preparation method according to claim 1, characterized in that: In step (1), the ball milling conditions are: filling rate 70-90%, rotation speed 1500-3000r / min, and ball milling time 30-60min.

6. The preparation method according to claim 1, characterized in that: In step (2), the cooking conditions are: the pressure is maintained at 0.25±2Mpa, the temperature is maintained at 120±5°C, and the cooking time is 3-8h.

7. The preparation method according to claim 1, characterized in that: In step (3), the mass ratio of the lipase to the alkaline protease is 1:2-5; the mass of the first complex enzyme added is 1-3% of the raw material 2.

8. The preparation method according to claim 1, characterized in that: In step (4), the mass ratio of papain, bromelain and carboxypeptidase is 1:0.5-2:1-5, and the mass of the second composite enzyme added is 0.5-2% of the raw material 2.

9. Calf thymosin prepared by the preparation method according to any one of claims 1 to 8.

10. Use of the calf thymosin peptide prepared by the preparation method according to any one of claims 1 to 8 in the preparation of products for improving immunity.

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