Liquid phase synthesis method of bursin

Abstract

The invention relates to the technical field of polypeptide synthesis, and particularly discloses a liquid-phase synthesis method of bursin tripeptide, which comprises the following steps: protection of lysine, preparation of a THF solution of Boc-Lys (Boc)-His-OMe, preparation of an ethyl acetate solution of Boc-Lys (Boc)-His-OH, preparation of an ethyl acetate solution of Boc-Lys (Boc)-His-Gly-NH2, and synthesis of bursin tripeptide. The whole synthesis process can be carried out in the same reactor, intermediate materials are prevented from being transferred among different reactors, the synthesis operation is simplified, meanwhile, the cost of raw materials and reagents is low, the reaction condition is mild, the route is stable, and the bursin tripeptide product meeting the purity requirement can be directly synthesized; the cost required by purification and the product loss in the purification process are reduced, and the synthesis period is greatly shortened.

Description

Technical Field

[0001] The invention relates to the technical field of polypeptide synthesis, and in particular to a liquid phase synthesis method of bursin tripeptide. Background Art

[0002] Tripeptide of bursin is a bioactive substance existing in the bursa of Fabricius of birds. It has a unique composition structure. It has been basically confirmed that it is closely related to immune function and has been widely used in the prevention and treatment of some diseases of poultry. The existing synthesis methods mainly include biosynthesis using enzymes, solid phase synthesis using resins, and liquid phase synthesis. Among them, biosynthesis and solid phase synthesis methods have high raw material costs and long cycles due to their process characteristics (mainly manifested in the use of enzymes and resins). At the same time, the synthesis operation is cumbersome, the reaction conditions are strict, the crude product yield is low, and the purity is low. Purification is required to obtain a bursin tripeptide product that meets the required purity requirements. Although the existing liquid phase synthesis method avoids the use of enzymes or resins and reduces part of the synthesis cost, it still has problems such as cumbersome operation, long synthesis cycle, and the need for purification, and the cost of raw materials is still high.

[0003] For the bursin tripeptide, patent CN105646648A chooses to use the solid phase chemical synthesis method of resin for synthesis, reacting the resin with a mixed solution of protected amino acids and condensation agents to complete the condensation of Gly, His and Lys in sequence, but the cost of the resin used in the solid phase synthesis is relatively high, and the raw materials need to be greatly excessive during the reaction, further increasing the cost; in order to remove the residual raw materials, a large amount of organic solvents such as DMF need to be used for washing after each condensation reaction; at the same time, the product obtained after solid phase synthesis cleavage is trifluoroacetate, which cannot be used directly, and the purity of the product does not meet the requirements, and further salt conversion purification is required. The purified product cannot be directly extracted from the aqueous solution and needs to be freeze-dried, which is cumbersome and requires high equipment requirements.

[0004] Patent CN1696151A selects liquid phase chemical synthesis for the synthesis of cystin tripeptide, but the synthesis method is cumbersome and requires multiple transfers, drying, and evaporation of the reaction solution; at the same time, the reaction time is long, a single condensation reaction takes several hours, and the reaction solution needs more than ten hours of drying time after post-treatment; the reaction conditions are strict, especially in the control of reaction temperature, there is a low temperature operation of -5°C, and the maximum temperature needs to be heated to 80°C for solvent reflux reaction, which increases energy consumption and the risk of large-scale preparation.

[0005] Patent CN101045745A provides two synthesis methods for bursin tripeptide. One is an azide-active ester synthesis method, in which active ester is selected for synthesis in the condensation reaction of lysine and histidine. The operation is cumbersome and the temperature needs to be controlled between -10°C and -2°C. The reaction takes several hours. A large amount of DCU solid by-product will precipitate during the reaction. Although the DCU by-product can be removed by filtration, a small amount of DCU will still remain in the solution and needs to be completely removed after multiple treatments. The azide method is selected for synthesis in the condensation of lysine-histidine and glycinamide. Azide compounds have explosive properties due to their chemical structure characteristics, and the reaction conditions are very harsh. They are extremely dangerous and uncontrollable in large-scale preparation. Another method is the trypsin enzymatic synthesis method, which uses enzyme biosynthesis. The cost of enzyme use is high and the environmental requirements are extremely high. Environmental changes can easily lead to enzyme inactivation. The enzyme synthesis method has a long synthesis cycle and the patent does not give the synthesis time. The synthesis yield is low, and the solution system after synthesis is complex with a variety of different components. Purification is required to obtain a usable cystin tripeptide product. Summary of the invention

[0006] The purpose of the present invention is to provide a continuous, low-cost, and efficient liquid phase synthesis method. The entire synthesis process can be carried out in the same reactor, avoiding the transfer of intermediate materials between different reactors, simplifying the synthesis operation, and at the same time, the cost of raw materials and reagents is low, and the reaction conditions are mild and the route is stable. A cystin tripeptide product that meets the purity requirements can be directly synthesized, reducing the cost required for purification and the product loss in the purification process, and the synthesis cycle is greatly shortened.

[0007] To achieve the above object, the technical solution adopted by the present invention is: The liquid phase synthesis method of bursin tripeptide specifically comprises the following steps: S1. Protection of lysine: Dissolve L-lysine in water, place in a water bath and stir, and add sodium hydroxide in batches, keeping the solution temperature at 25℃~30℃. After the sodium hydroxide is dissolved, add Boc anhydride dissolved in THF (tetrahydrofuran) dropwise, control the temperature between 25℃~30℃ to react for 10~30 minutes, then add hydrochloric acid dropwise until the pH is 2~3, add sodium chloride until the aqueous solution is saturated, continue stirring, and then separate the liquids to take the upper organic phase to obtain a THF solution of Boc-Lys(Boc)-OH with protected amino groups; S2. Preparation of THF solution of Boc-Lys(Boc)-His-OMe: L-histidine methyl ester dihydrochloride is added to the above-mentioned THF solution of Boc-Lys(Boc)-OH and stirred, and then triethylamine is added. After stirring evenly, TBTU (2-(1H-benzotriazol L-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate) dissolved in DMF (N,N-dimethylformamide) is added dropwise, and the temperature is controlled between 25 and 30°C to react for 10 to 20 minutes, and then sodium chloride aqueous solution and THF are added, stirring is continued, and then the upper organic phase is separated; then the organic phase is added with sodium chloride aqueous solution and hydrochloric acid to pH=5 to 6 and washed twice, and then the organic phase is added with sodium chloride aqueous solution and sodium carbonate to pH=9 to 10 and washed twice, and then washed once with saturated sodium chloride aqueous solution, the upper organic phase is separated, and the organic phase is concentrated to obtain a THF solution of Boc-Lys(Boc)-His-OMe; S3. Preparation of ethyl acetate solution of Boc-Lys(Boc)-His-OH: Add sodium hydroxide dissolved in water to the THF solution of Boc-Lys(Boc)-His-OMe, react at room temperature for 10-30 minutes, then add hydrochloric acid dropwise to pH=5-6, add sodium chloride until the aqueous solution is saturated, and add an additional saturated sodium chloride aqueous solution 2 times the volume of the THF solution, add ethyl acetate 3 times the volume of the THF solution, continue stirring, then separate the upper organic phase, and concentrate the organic phase to obtain an ethyl acetate solution of Boc-Lys(Boc)-His-OH; S4, ethyl acetate solution of Boc-Lys(Boc)-His-Gly-NH2: Glycinamide hydrochloride is added to the ethyl acetate solution of Boc-Lys(Boc)-His-OH and stirred, and then triethylamine is added. After stirring evenly, TBTU dissolved in DMF is added dropwise, and the temperature is controlled between 25 and 30°C to react for 10 to 20 minutes. Then, sodium chloride aqueous solution and ethyl acetate are added, and stirring is continued. Then, the upper organic phase is separated and then sodium chloride aqueous solution is added to the organic phase and hydrochloric acid is added to the pH value of 5 to 6 and washed twice. Then, sodium chloride aqueous solution is added to the organic phase and sodium carbonate is added to the organic phase to pH value of 9 to 10 and washed twice, and then washed once with saturated sodium chloride aqueous solution, and the upper organic phase is separated and then taken to obtain an ethyl acetate solution of Boc-Lys(Boc)-His-Gly-NH2; S5. Synthesis of bursa tripeptide: Triisopropylsilane was added to the ethyl acetate solution of Boc-Lys(Boc)-His-Gly-NH2, and after stirring evenly, an ethyl acetate solution containing HCl was added, and the mixture was stirred at room temperature for 10 to 30 minutes. Then, methyl tert-butyl ether was added to the reaction solution, and the mixture was stirred for crystallization for 60 to 90 minutes, and then filtered to obtain a white solid. The white solid was dried to remove the residual solvent on the surface, and finally the desired pure Lys-His-Gly-NH2·HCl of the tripeptide cysteine ​​with a purity of more than 95% was obtained.

[0008] Preferably, the molar ratio of L-lysine, Boc anhydride and sodium hydroxide used in step S1 is 1:2:3, and the volume ratio of water and THF is 5:3.

[0009] Preferably, the molar ratio of L-histidine methyl ester dihydrochloride used in step S2 to L-lysine used in step S1 is 1:1.

[0010] Preferably, the amount of TBTU used in step S2 is: 1.1eq≥equivalent≥1.05eq; the amount of triethylamine used is: 3.1eq≥equivalent≥3.05eq; the amount of sodium chloride aqueous solution added each time is 3-4 times the volume of DMF, the concentration of the first sodium chloride aqueous solution is 20%, the concentration of the second sodium chloride aqueous solution is 15%, and the concentration of the third sodium chloride aqueous solution is 8%; the amount of THF added is 3-4 times the volume of DMF.

[0011] Preferably, the amount of sodium hydroxide used in step S3 is: 4eq≥equivalent≥3eq; the volume of water used to dissolve the sodium hydroxide is the same as the volume of the THF solution after concentration in step S2.

[0012] Preferably, the molar ratio of glycineamide hydrochloride used in step S4 to L-lysine used in step S1 is 1:1.

[0013] Preferably, the amount of TBTU used in step S4 is: 1.1eq≥equivalent≥1.05eq; the amount of triethylamine used is: 3.1eq≥equivalent≥3.05eq; the amount of sodium chloride aqueous solution added each time is 3-4 times the volume of DMF, the concentration of the first sodium chloride aqueous solution is 20%, the concentration of the second sodium chloride aqueous solution is 15%, and the concentration of the third sodium chloride aqueous solution is 8%; the amount of ethyl acetate added is 2-3 times the volume of DMF.

[0014] Preferably, in step S5, the molar amount of HCl is 7.5-10 times the molar amount of the Boc protecting group to be removed; in order to reduce the generation of impurities during the removal of the Boc protecting group and thus improve the purity of the product, the molar amount of triisopropylsilane added is 2-3 times the molar amount of the Boc protecting group to be removed.

[0015] Preferably, the liquid separation in step S1 and step S2 is carried out from the upper organic phase, the upper layer is the THF organic phase during liquid separation, the aqueous phase is discharged from the lower discharge port, and the THF solution is retained in the reaction container to continue the subsequent step reaction.

[0016] Preferably, the liquid separation in step S3 and step S4 is taken from the upper organic phase, the upper layer is the ethyl acetate organic phase during liquid separation, the aqueous phase is discharged from the lower discharge port, and the ethyl acetate solution is retained in the reaction container to continue the subsequent step reaction.

[0017] Compared with the prior art, the present invention has the following beneficial effects: In the process of post-treatment with sodium chloride solution, each intermediate organic solution prepared in the synthesis process of the present invention has an organic phase in the upper layer during liquid separation, and an aqueous phase is separated from the bottom, and the required organic phase is retained in the reactor. The organic solution between each step does not need to be transferred out of the reactor, and continuous synthesis is carried out in the same reactor until the cysteine ​​tripeptide is finally obtained. The reaction conditions are mild and the route is stable. A cysteine ​​tripeptide product that meets the purity requirements can be directly synthesized, the cost required for purification and the product loss in the purification process are reduced, the synthesis cycle is greatly shortened, and the entire synthesis can be completed in one day.

[0018] In the present invention, in order to ensure the continuity of the reaction and to carry out the reaction in the same reactor, the solution prepared in steps S1 and S2 is a THF solution; in order to reduce the content of residual water in the prepared solution and thus reduce the adverse effect of water on the amide condensation reaction, the water solubility of ethyl acetate is lower than that of THF, and in steps S4 and S5 where no sodium hydroxide is used, ethyl acetate is selected as the reaction solvent and prepared into an ethyl acetate solution, and the solvent conversion is completed in step S3. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 This is the ELSD spectrum of Boc-Lys(Boc)-OH of the present invention.

[0020] Figure 2 The HPLC spectrum of Boc-Lys(Boc)-His-OMe of the present invention.

[0021] Figure 3 The HPLC spectrum of Boc-Lys(Boc)-His-OH of the present invention.

[0022] Figure 4 The HPLC spectrum of Boc-Lys(Boc)-His-Gly-NH2 of the present invention.

[0023] Figure 5 The HPLC spectrum of Lys-His-Gly-NH2•HCl of the present invention. DETAILED DESCRIPTION

[0024] In order to make the above features and advantages of the present invention more obvious and easy to understand, embodiments are given below with reference to the accompanying drawings for detailed description as follows.

[0025] This embodiment provides a liquid phase synthesis method of bursin tripeptide, comprising the following steps: protection of lysine, preparation of THF solution of Boc-Lys(Boc)-His-OMe, preparation of ethyl acetate solution of Boc-Lys(Boc)-His-OH, ethyl acetate solution of Boc-Lys(Boc)-His-Gly-NH2, and synthesis of bursin tripeptide. Specifically: S1. Protection of lysine: 0.020 mol L-lysine was dissolved in 25 mL of water, stirred in a water bath and 0.060 mol of sodium hydroxide was added in batches, the solution temperature was maintained at 30°C, after the sodium hydroxide was dissolved, 0.040 mol of Boc anhydride dissolved in 15 mL of THF was added dropwise, the temperature was controlled between 25-30°C for 30 minutes, and then hydrochloric acid was added dropwise until pH = 3 (in some other examples, pH can also be 2 or a point between 2 and 3), sodium chloride was added until the aqueous solution was saturated, stirring was continued for 5 minutes, and then the upper organic phase was separated. The upper layer during separation was the THF organic phase, and the aqueous phase was discharged from the lower discharge port. The THF solution was retained in the reaction vessel to continue the subsequent steps to obtain a THF solution of Boc-Lys(Boc)-OH with protected amino groups. The purity test results of ELSD (evaporative light scattering detector) are as follows Figure 1 .

[0026] S2. Preparation of THF solution of Boc-Lys(Boc)-His-OMe: 0.020 mol L-histidine methyl ester dihydrochloride was added to the THF solution of Boc-Lys(Boc)-OH and stirred, then 0.0615 mol triethylamine was added, and after stirring evenly, 0.022 mol TBTU dissolved in 25 mL DMF was added dropwise, and the temperature was controlled between 25-30°C for 20 minutes, and then 75 mL 20% aqueous sodium chloride solution and 75 mL THF, continue stirring for 3 minutes, then separate and take the upper organic phase, the upper layer is THF organic phase during separation, the aqueous phase is discharged from the lower discharge port, and the THF solution is retained in the reaction vessel to continue the subsequent steps of the reaction; then add 75mLmL of 15% sodium chloride aqueous solution to the organic phase and add hydrochloric acid to pH=5 (in some other examples, the pH can also be 6 or a point between 5 and 6) and wash twice, then add 75mLmL of 8% sodium chloride aqueous solution to the organic phase and add sodium carbonate to pH=10 (in some other examples, the pH can also be 9 or a point between 9 and 10) and wash twice, then wash once with 75mLmL of saturated sodium chloride aqueous solution, separate and take the upper organic phase, the upper layer is THF organic phase during separation, the aqueous phase is discharged from the lower discharge port, and the THF solution is retained in the reaction vessel to continue the subsequent steps of the reaction; the organic phase is concentrated to about 25mL remaining to obtain a THF solution of Boc-Lys(Boc)-His-OMe, and the HPLC (high performance liquid chromatography) purity test results are as follows Figure 2 .

[0027] S3. Preparation of ethyl acetate solution of Boc-Lys(Boc)-His-OH: add 0.070 mol of sodium hydroxide dissolved in 25 mL of water to the THF solution of Boc-Lys(Boc)-His-OMe, react at room temperature for 25 minutes, then add hydrochloric acid dropwise until pH = 5 (in some other examples, pH can also be 6 or a point between 5 and 6), add sodium chloride until the aqueous solution is saturated, and add an additional 50 mL of saturated sodium chloride aqueous solution, add 75 mL of ethyl acetate, continue stirring for 3 minutes, then separate the liquid and take the upper organic phase. During separation, the upper layer is the ethyl acetate organic phase. The aqueous phase is discharged from the lower discharge port, and the ethyl acetate solution is retained in the reaction vessel to continue the subsequent step reaction; the organic phase is concentrated to about 25 mL remaining to obtain the ethyl acetate solution of Boc-Lys(Boc)-His-OH. The HPLC purity test results are as follows: Figure 3 .

[0028] S4, ethyl acetate solution of Boc-Lys(Boc)-His-Gly-NH2: add 0.020 mol of glycineamide hydrochloride to the above ethyl acetate solution of Boc-Lys(Boc)-His-OH and stir, then add 0.0615 mol of triethylamine, stir evenly and dropwise add 0.022 mol of TBTU dissolved in 25 mL of DMF, control the temperature between 25-30°C to react for 20 minutes, then add 75 mL of 20% sodium chloride aqueous solution and 75 mL of ethyl acetate, continue stirring for 5 minutes, then separate the liquids and take the upper organic phase, the upper layer during separation is the ethyl acetate organic phase, the aqueous phase is discharged from the lower discharge port, and the ethyl acetate solution is retained in the reaction vessel to continue the subsequent step reaction; then add 75 mL of 15% sodium chloride aqueous solution to the organic phase and add hydrochloric acid to pH = 5 (in some other examples, the pH can also be 6 or a point between 5 and 6), wash twice, then add 75 mL of 8% sodium chloride aqueous solution and sodium carbonate was added to pH = 10 (in some other examples, pH can also be 9 or a point between 9 and 10) and washed twice, and then washed once with 75 mL of saturated sodium chloride aqueous solution, and the upper organic phase was separated. The upper layer during separation was ethyl acetate organic phase, and the aqueous phase was discharged from the lower discharge port. The ethyl acetate solution was retained in the reaction vessel to continue the subsequent step reaction; the organic phase was concentrated to about 25 mL remaining to obtain an ethyl acetate solution of Boc-Lys(Boc)-His-Gly-NH2. The HPLC purity test results are as follows Figure 4 .

[0029] S5. Synthesis of bursin tripeptide: add 0.100 mol of triisopropylsilane to the ethyl acetate solution of Boc-Lys(Boc)-His-Gly-NH2, stir evenly, add 75 ml of 4 mol / L HCl in ethyl acetate, stir at room temperature for 30 minutes, then add 100 mL of methyl tert-butyl ether to the reaction solution, stir and crystallize for 60 minutes, then filter to obtain a white solid, place the white solid in an oven at 35°C to dry to remove the residual solvent on the surface, and after drying, obtain 7.36 g of pure bursin tripeptide Lys-His-Gly-NH2•HCl, with a yield of 81.95% and a purity of 96.73%. The HPLC purity test results are as follows: Figure 5 .

[0030] The above shows and describes the basic principles, main features and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The above embodiments and descriptions are only for illustrating the principles of the present invention. Without departing from the spirit and scope of the present invention, the present invention may have various changes and improvements, which shall fall within the scope of the present invention to be protected. The scope of protection of the present invention shall be defined by the attached claims and their equivalents.

Claims

1. A liquid phase synthesis method of bursin tripeptide, characterized in that: The specific steps include: S1. Protection of lysine: Dissolve L-lysine in water, place in a water bath and stir, and add sodium hydroxide in batches, keeping the solution temperature at 25°C~30°C. After the sodium hydroxide is dissolved, add Boc anhydride dissolved in THF dropwise, control the temperature between 25°C~30°C to react for 10~30 minutes, then add hydrochloric acid dropwise until the pH is 2~3, add sodium chloride until the aqueous solution is saturated, continue stirring, and then separate and take the upper organic phase to obtain a THF solution of Boc-Lys(Boc)-OH with protected amino groups; S2. Preparation of THF solution of Boc-Lys(Boc)-His-OMe: L-histidine methyl ester dihydrochloride is added to the above-mentioned Boc-Lys(Boc)-OH THF solution and stirred, and then triethylamine is added. After stirring evenly, TBTU dissolved in DMF is added dropwise, and the temperature is controlled between 25 and 30° C. to react for 10 to 20 minutes, and then sodium chloride aqueous solution and THF are added, stirring is continued, and then the upper organic phase is separated; then the organic phase is added with sodium chloride aqueous solution and hydrochloric acid to pH=5 to 6 and washed twice, and then the organic phase is added with sodium chloride aqueous solution and sodium carbonate to pH=9 to 10 and washed twice, and then washed once with saturated sodium chloride aqueous solution, the upper organic phase is separated, and the organic phase is concentrated to obtain a THF solution of Boc-Lys(Boc)-His-OMe; S3. Preparation of ethyl acetate solution of Boc-Lys(Boc)-His-OH: Add sodium hydroxide dissolved in water to the THF solution of Boc-Lys(Boc)-His-OMe, react at room temperature for 10-30 minutes, then add hydrochloric acid dropwise to pH=5-6, add sodium chloride until the aqueous solution is saturated, and add an additional saturated sodium chloride aqueous solution 2 times the volume of the THF solution, add ethyl acetate 3 times the volume of the THF solution, continue stirring, then separate the upper organic phase, and concentrate the organic phase to obtain an ethyl acetate solution of Boc-Lys(Boc)-His-OH; S4, ethyl acetate solution of Boc-Lys(Boc)-His-Gly-NH2: Glycinamide hydrochloride is added to the ethyl acetate solution of Boc-Lys(Boc)-His-OH and stirred, and then triethylamine is added. After stirring evenly, TBTU dissolved in DMF is added dropwise, and the temperature is controlled between 25 and 30°C to react for 10 to 20 minutes. Then, sodium chloride aqueous solution and ethyl acetate are added, and stirring is continued. Then, the upper organic phase is separated and then sodium chloride aqueous solution is added to the organic phase and hydrochloric acid is added to the pH value of 5 to 6 and washed twice. Then, sodium chloride aqueous solution is added to the organic phase and sodium carbonate is added to the organic phase to pH value of 9 to 10 and washed twice, and then washed once with saturated sodium chloride aqueous solution, and the upper organic phase is separated and then taken to obtain an ethyl acetate solution of Boc-Lys(Boc)-His-Gly-NH2; S5. Synthesis of bursa tripeptide: Triisopropylsilane was added to the ethyl acetate solution of Boc-Lys(Boc)-His-Gly-NH2, and after stirring evenly, an ethyl acetate solution containing HCl was added, and the mixture was stirred at room temperature for 10 to 30 minutes. Then, methyl tert-butyl ether was added to the reaction solution, and the mixture was stirred for crystallization for 60 to 90 minutes, and then filtered to obtain a white solid. The white solid was dried to remove the residual solvent on the surface, and finally the desired pure Lys-His-Gly-NH2·HCl of the tripeptide cysteine ​​with a purity of more than 95% was obtained.

2. The liquid phase synthesis method of the bursin tripeptide according to claim 1, characterized in that: The molar ratio of L-lysine, Boc anhydride and sodium hydroxide used in step S1 is 1:2:3, and the volume ratio of water and THF is 5:

3.

3. The liquid phase synthesis method of the bursin tripeptide according to claim 1, characterized in that: The molar ratio of L-histidine methyl ester dihydrochloride used in step S2 to L-lysine used in step S1 is 1:

1.

4. The liquid phase synthesis method of the bursin tripeptide according to claim 1, characterized in that: The amount of TBTU used in the step S2 is: 1.1eq≥equivalent≥1.05eq; the amount of triethylamine used is: 3.1eq≥equivalent≥3.05eq; the amount of sodium chloride aqueous solution added each time is 3-4 times the volume of DMF, the concentration of the first sodium chloride aqueous solution is 20%, the concentration of the second sodium chloride aqueous solution is 15%, and the concentration of the third sodium chloride aqueous solution is 8%; the amount of THF added is 3-4 times the volume of DMF.

5. The liquid phase synthesis method of bursin tripeptide according to claim 1, characterized in that: The amount of sodium hydroxide used in step S3 is: 4eq≥equivalent≥3eq; the volume of water used to dissolve the sodium hydroxide is the same as the volume of the THF solution after concentration in step S2.

6. The liquid phase synthesis method of bursin tripeptide according to claim 1, characterized in that: The molar ratio of glycine amide hydrochloride used in step S4 to L-lysine used in step S1 is 1:

1.

7. The liquid phase synthesis method of bursin tripeptide according to claim 1, characterized in that: The amount of TBTU used in the step S4 is: 1.1eq≥equivalent≥1.05eq; the amount of triethylamine used is: 3.1eq≥equivalent≥3.05eq; the amount of sodium chloride aqueous solution added each time is 3-4 times the volume of DMF, the concentration of the first sodium chloride aqueous solution is 20%, the concentration of the second sodium chloride aqueous solution is 15%, and the concentration of the third sodium chloride aqueous solution is 8%; the amount of ethyl acetate added is 2-3 times the volume of DMF.

8. The liquid phase synthesis method of bursin tripeptide according to claim 1, characterized in that: The step S5 is to remove the Boc protecting group, and the molar amount of HCl is 7.5-10 times the molar amount of the Boc protecting group to be removed; the molar amount of triisopropylsilane added is 2-3 times the molar amount of the Boc protecting group to be removed.

9. The liquid phase synthesis method of bursin tripeptide according to claim 1, characterized in that: The liquid separation in step S1 and step S2 is carried out by taking the upper organic phase, wherein the upper layer is the THF organic phase during liquid separation, and the aqueous phase is discharged from the lower discharge port, and the THF solution is retained in the reaction container to continue the subsequent step reaction.

10. The liquid phase synthesis method of bursin tripeptide according to claim 1, characterized in that: The liquid separation in step S3 and step S4 is carried out by taking the upper organic phase, wherein the upper layer is the ethyl acetate organic phase, and the aqueous phase is discharged from the lower discharge port, and the ethyl acetate solution is retained in the reaction container to continue the subsequent step reaction.

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