CN113637064B

CN113637064B - Synthetic method of somalupeptide

Info

Publication number: CN113637064B
Application number: CN202010393322.2A
Authority: CN
Inventors: 张利香; 付玉清; 姚林; 朱亮平; 李文静
Original assignee: Shenzhen Jx Bio Pharmaceutical Co ltd
Current assignee: Shenzhen Jx Bio Pharmaceutical Co ltd
Priority date: 2020-05-11
Filing date: 2020-05-11
Publication date: 2023-11-28
Anticipated expiration: 2040-05-11
Also published as: CN113637064A

Abstract

The invention discloses a synthesis method of a somalupeptide, which comprises the following steps: the S1-S3 fragment and the S4-S6 fragment which are fully protected by the somalupeptide are respectively synthesized and used for synthesizing the somalupeptide resin, and the somalupeptide is obtained through cracking and purifying. The method also comprises the step of synthesizing the lysine with the side chain group at the 20 th position of the sorulon, and the segment is favorable for completing the coupling of the sorulon main chain, so that the purity and the yield of a sorulon crude product are greatly improved, and the synthesis cost is reduced. In the method, the fully protected S1-S3 fragment and the fully protected S4-S6 fragment of the somalundum are prepared, and are used as key starting materials for solid-phase synthesis of the somalundum, so that the production of D-His, D-Glu, D-Thr, D-Phe racemization impurities and +Gly impurities is greatly reduced, the difficulty of crude purification is obviously reduced, the purity and yield of the somalundum are greatly improved, the synthesis cost is reduced, and the industrial mass production is facilitated.

Description

Synthetic method of somalupeptide

Technical Field

The invention relates to the field of polypeptide synthesis, in particular to a method for preparing a somalupeptide.

Technical Field

Glucagon-like peptide-1 (GLP-1) is a peptide hormone secreted by human intestinal L cells, and has effects of promoting insulin secretion, inhibiting glucagon secretion, and lowering blood sugar concentration, and can be used for treating type II diabetes. However, natural GLP-1 is unstable in vivo and is susceptible to rapid degradation by dipeptidyl peptidase-IV (DPP-IV).

Somamunotide, named Semaglutide, is a novel long-acting glucagon-like peptide-1 (GLP-1) analog developed and produced by Daneno and Norde corporation for the treatment of type II diabetes. The somalunin has the effects of reducing blood sugar, losing weight and protecting cardiovascular system, and is approved by FDA in 12 months of 2017. After the Lys side chain of the somalundum is modified by PEG, glu and octadecadicarboxylic acid, the hydrophilicity is greatly improved, and the binding force with albumin is enhanced; meanwhile, after Ala at the 2 nd position of the N end is mutated into Aib, the inactivation caused by DPP-IV enzymolysis is effectively avoided, the half life reaches 40h, and patients only need to inject once a week. The CAS number of the somalundum is 910463-68-2, the molecular formula is C187H291N45O59, the molecular weight is 4113.64g/mol, and the peptide sequence is:

H-His-Aib-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Val-Ser-Ser-Tyr-Leu-Glu-Gly-Gln-Ala-Ala-Lys(AEEA-AEEA-γ-Glu-Octadecanedioic)-Glu-Phe-Ile-Ala-Trp-Leu-Val-Arg-Gly-Arg-Gly-OH。

the methods for preparing somalundin reported so far are roughly divided into two categories: one is to directly access Lys containing side chain as fragment into the main chain of the somalupeptide to complete synthesis, patent CN104356224A discloses a method of using liquid phase method to synthesize epsilon-NH of Lys ₂ A method for preparing the somalundin by grafting a side chain and then gradually condensing amino acid on resin. The other is to finish the coupling of the main chain and the side chain of the cable-marlutide one by one, and patent CN 201511027176 discloses that the cable-marlutide linear peptide is synthesized gradually in a solid phase, a side chain modification group is synthesized, a protecting group of Lys is removed, the side chain modification group is coupled, and finally the polypeptide product is obtained by cleavage. Because the sequence of the somalupeptide is longer and has more hydrophobic amino acids, the somalupeptide is easy to form folding when synthesized by adopting an amino acid gradual condensation method, the shrinkage of resin is serious, the reaction time is prolonged, and more impurities with very similar properties to products, such as [ D-His ] are generated in the crude peptide]Is a racemic impurity of (2)

H-D-His-Aib-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Val-Ser-Ser-Tyr-Leu-Glu-Gly-Gln-Ala-Ala-Lys(AEEA-AEEA-γ-Glu-Octadecane-dioic)-Glu-Phe-Ile-Ala-Trp-Leu-Val-Arg-Gly-Arg-Gly-OH；+Gly

Impurity(s)

H-His-Aib-Glu-Gly-Gly-Thr-Phe-Thr-Ser-Asp-Val-Ser-Ser-Tyr-Leu-Glu-Gly-Gln-Ala-Ala-Lys(AEEA-AEEA-γ-Glu-Octadecanedioic)-Glu-Phe-Ile-Ala-Trp-Leu-Val-Arg-Gly-Arg-Gly-OH；D-Phe

Impurity(s)

H-His-Aib-Glu-Gly-Thr-D-Phe-Thr-Ser-Asp-Val-Ser-Ser-Tyr-Leu-Glu-Gly-Gln-Ala-Ala-Lys(Octadecanedioicacidmono-tert-butyl

ester-gamma-Glu-PEG-PEG) -Glu-Phe-Ile-Ala-Trp-Leu-Val-Arg-Gly-Arg-Gly-OH; the [ D-His ] racemization impurity is similar to the physicochemical property of the target peptide of the somalundum, so that the separation and purification difficulty of the somalundum product is greatly increased, and the product yield is greatly reduced. Therefore, there is an urgent need for a method for synthesizing somalundin with high purity and yield and low synthesis cost.

Disclosure of Invention

The invention provides a synthesis method of the simaroubm peptide with high yield and high purity, which aims to solve the problems of more impurities, low purity and yield, high cost, complex operation steps, unfavorable industrial production and the like in the existing synthesis process. The strategy of adding tripeptide and tripeptide fragments is adopted to reduce the generation of D-His racemization impurity, +Gly impurity, D-Thr and D-Phe impurity, and has considerable economic and applicable value and wide application prospect.

In order to achieve the aim of the invention, the invention provides a synthesis method of the somalundum, which is used for respectively synthesizing the somalundum full protection S1-S3 fragment and the S4-S6 fragment, and is applied to the synthesis of the somalundum full protection peptide resin, and the somalundum is obtained through cracking and purifying.

It should be noted that in the present invention, S1 is the first amino acid at the N-terminal in the somalupeptide peptide sequence, and similarly S2, S3, S4, S5, S6, S18, S19, S20 are the 2 nd, 3 rd, 4 th, 5 th, 6 th, 18 th, 19 th, 20 th amino acids at the N-terminal, respectively, and so on.

Preferably, the fully protected S1-S3 fragment is R1-His (R2) -Aib-Glu (OR) ₃ ) The full-protection S4-S6 fragment is R4-Gly-Thr (R5) -Phe-OH, wherein

R ₁ Selected from the group consisting of Fmoc, dde, alloc, boc, trt, dmb, mmt, mtt,

R ₂ selected from the group consisting of Fmoc, dde, alloc, boc, trt, dmb, mmt, mtt,

R ₃ is selected from the group consisting of tBu and Bzl,

R ₄ selected from the group consisting of Fmoc,

R ₅ selected from tBu and Bzl.

More preferably, R1 is selected from: boc or Fmoc, R2 is selected from: trt or Boc, R3 is selected from tBu, R4 is selected from Fmoc, and R5 is selected from tBu.

In some embodiments R ₁ Is Boc, R ₂ Is Trt, R ₃ Is tBu, R ₄ Fmoc, R ₅ Is tBu, namely: the S1-S3 fragment is Boc-His (Trt) -Aib-Glu (OtBu) -OH, and the S4-S6 fragment is Fmoc-Gly-Thr (tBu) -Phe-OH. The adoption of the full-protection S1-S3 fragment Boc-His (Trt) -Aib-Glu (OtBu) -OH can effectively reduce the generation of D-His racemization impurities; the adoption of full protection, the S4-S6 fragment Fmoc-Gly-Thr (tBu) -Phe-OH can effectively reduce the generation of +Gly impurity, D-Thr and D-Phe impurity, and the use of the two fragments can obviously improve the yield and purity of the crude peptide of the somalundin.

The applicant has unexpectedly found that in the preparation of the somalundum peptide, the monomer Fmoc-Lys (AEEa-AEEa-gamma-Glu (OtBu) -Octadecanantioic) -OH is adopted at the 20 th position, so that a peptide intermediate can be easily inserted into SPPS, an amino terminal amino acid can be more easily reacted with the SPPS, mismatched peptide impurities (such as amino acid missing peptide and amino acid superfluous peptide) and racemic peptide impurities can be obviously inhibited/reduced, and the yield and purity of the crude somalundum peptide are obviously improved.

In some embodiments, S20 is: fmoc-Lys (AEEA-AEEA-gamma-Glu (OtBu) -Octadecananioic) -OH, S1-S3 fragments are: the Boc-His (Trt) -Aib-Glu (OtBu) -OH, S4-S6 fragment is: fmoc-Gly-Thr (tBu) -Phe-OH.

According to the invention, the solid-phase synthesis method combining the fragment and the stepwise synthesis is adopted to prepare the S1-S3 fragment and the S4-S6 fragment of the somalupeptide, and the fragments are used as key starting materials for the solid-phase synthesis of the somalupeptide, so that the generation of D-His, D-Glu, D-Thr, D-Phe racemization impurities and +Gly impurities is greatly reduced, the difficulty of crude product purification is obviously reduced, the purity and yield of the somalupeptide are greatly improved, the synthesis cost is reduced, and the industrial mass production is facilitated.

Drawings

FIG. 1 is an HPLC chromatogram of crude peptide of somalupeptide prepared in example 8

FIG. 2 is an HPLC chromatogram of the somalupeptide refined peptide prepared in example 11

Detailed Description

The foregoing of the invention will be described in further detail with reference to specific embodiments. It should not be understood that the scope of the above subject matter of the present invention is limited to the following examples only. All techniques implemented based on the above description of the invention are within the scope of the invention.

The meanings of the abbreviations used in the present invention are listed in the following table:

EXAMPLE 1 preparation of Boc-His (Trt) -Aib-Glu (OtBu) -OH

A. 150g of 2-CTC resin with substitution degree of 1.06mmol/g is added into a reaction kettle, 500ml of dichloromethane is added, after mixing for 2min, dichloromethane is filtered off, 500ml of dichloromethane is added again, after mixing for 40min, dichloromethane is filtered off, finally 500ml of dichloromethane is added again, after mixing for 2min, dichloromethane is filtered off, and the resin is ready for use.

B. 135.30g of Fmoc-Glu (OtBu) -OH was weighed into a beaker, 500ml of DMF and 78.83ml of DIEA were added, the solution was stirred at 0-10℃for 5min and then poured into the CTC resin obtained in step A, and the mixture was reacted for 4h at 20-25 ℃. After the reaction is finished, DMF is filtered off. A mixed solution of 25ml of methanol and 250ml of DMF, a mixed solution of 50ml of DIEA and 250ml of DMF was added to the resin, and the mixing reaction was continued for 1h. After the reaction is finished, carrying out suction filtration, and washing the resin with DMF for 5 times, wherein 500ml of the resin is used each time; after washing, methanol is used for washing twice, and 500ml of methanol is used for each time; washing with dichloromethane for 2 times, each time with 500ml; finally, the resin was washed 3 times with 500ml portions of methanol until the resin was sufficiently dispersed.

C. And (3) drying the resin obtained in the step (B) in a vacuum drying oven at 20-30 ℃ for 4 hours until the weight is constant (twice weighing is carried out continuously, and the error is lower than 1%). After drying, 211.89g of Fmoc-Glu (OtBu) -CTC resin was obtained, and the degree of substitution was detected to be 0.75mmol/g.

D. The Fmoc-Glu (OtBu) -CTC resin obtained in step C was poured into a reaction vessel, swollen with 500ml DCM, mixed for 15min and then pumped down. 500ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 5min, followed by draining. DMF500ml was added and after mixing for 5min, the mixture was drained. 500ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 10min, followed by draining. DMF500ml was added and after mixing for 5min, the mixture was drained. Repeated DMF washing 5 times, 500ml each time, mixing 5min each time, and after washing 4 times, the filtrate was checked with PH test paper, which showed that PH was acceptable at 6.5-7.0.

E. 103.39g Fmoc-Aib-OH, 48.16g DIC and 51.57g HOBT were weighed in sequence in a clean 1L beaker, 500ml of DMF/DCM solution in a volume ratio of 1:1 was added, and the mixture was placed in ice water and stirred with a mechanical stirrer at 0-10℃for dissolution, and after the temperature was constant, the temperature was maintained and stirring was continued for 5min. Slowly adding the activating solution into a reaction kettle, and mixing and reacting for 2 hours at 20-25 ℃. After the reaction, the mixture was drained, 500ml of DMF was added thereto, and after 5 minutes of mixing, the mixture was drained. The DMF washes were repeated 5 times, 500ml each time, and 5min each time of mixing. Finally, detecting the obtained product as negative by ninhydrin to obtain Fmoc-Aib-Glu (OtBu) -CTC resin.

F. The amino acid Boc-His (Trt) -OH was coupled as described above for the deprotection method of step D and the coupling method of step E. Finally, washing with dichloromethane for 5 times, each time 500ml; after washing, methanol is used for washing twice, and 500ml of methanol is used for each time; washing with dichloromethane for 2 times, each time with 500ml; finally, the resin was washed 3 times with 500ml portions of methanol until the resin was sufficiently dispersed. The resin was dried in a vacuum oven at 20-30 ℃ for 4h to constant weight (twice weighing in succession, error lower than 1%). 245.31g of a fully protected Boc-His (Trt) -Aib-Glu (OtBu) -CTC resin was obtained.

G. The lysate ratio is TFE: DCM=1:4 (volume ratio), 20g of the full-protection peptide resin of the CTC resin obtained in the step F is added into 200mL of lysate at 15 ℃, the temperature is raised to 30 ℃, the reaction is continued for 3 hours by stirring, then the filtered resin is filtered by a sand core funnel, the filtered resin is washed by 100mL of DCM, the filtrate is combined after repeated operation for two times, the filtrate is concentrated under reduced pressure until the volume of the filtrate is 30% of the original volume, then the concentrated solution is slowly added into precooled 1L of isobutyl ether, the mixture is centrifuged for 5 times after sedimentation, 200mL of isobutyl ether is used each time, white solid powder is obtained, the mixture is dried by nitrogen gas firstly, then dried by a vacuum drying box for 10 hours, and 7.65g of full-protection Boc-His (Trt) -Aib-Glu (OtBu) -OH crude product is obtained after weighing.

EXAMPLE 2 preparation of Fmoc-His (Boc) -Aib-Glu (OBzl) -OH

A. 100g of 2-CTC resin with substitution degree of 1.12mmol/g is added into a reaction kettle, 300ml of dichloromethane is added, after mixing for 2min, dichloromethane is filtered off, 300ml of dichloromethane is added again, after mixing for 40min, dichloromethane is filtered off, finally 300ml of dichloromethane is added again, after mixing for 2min, dichloromethane is filtered off, and the resin is ready for use.

B. 102.93g of Fmoc-Glu (OBzl) -OH was weighed into a beaker, 300ml of DMF and 55.53ml of DIEA were added, the solution was stirred at 0-10℃for 5min and then poured into the CTC resin obtained in step A, and the mixture was reacted for 4h at 20-25 ℃. After the reaction is finished, DMF is filtered off. A mixed solution of 20ml of methanol and 150ml of DMF, a mixed solution of 30ml of DIEA and 150ml of DMF was added to the resin, and the mixing reaction was continued for 1h. After the reaction is finished, carrying out suction filtration, and washing the resin with DMF for 5 times, wherein 300ml of the resin is used each time; after washing, the mixture is washed twice with 300ml of methanol each time; washing with dichloromethane for 2 times, 300ml each time; finally, the resin was washed 3 times with 300ml portions of methanol until the resin was sufficiently dispersed.

C. And (3) drying the resin obtained in the step (B) in a vacuum drying oven at 20-30 ℃ for 4 hours until the weight is constant (twice weighing is carried out continuously, and the error is lower than 1%). After drying, 146.54g of Fmoc-Glu (OBzl) -CTC resin was obtained, which showed a degree of substitution of 0.76mmol/g.

D. The Fmoc-Glu (OBzl) -CTC resin obtained in step C was poured into the reaction vessel, swollen with 300ml DCM and mixed for 15min, and then pumped down. 500ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 5min, followed by draining. DMF (300 ml) was added thereto, and after mixing for 5 minutes, the mixture was drained. 300ml of 20% piperidine/DMF solution was added and mixed at 20-30℃for 10min, followed by draining. DMF (300 ml) was added thereto, and after mixing for 5 minutes, the mixture was drained. Repeated DMF washing for 5 times, 300ml each time, mixing for 5min each time, and after the 4 th washing, detecting the filtrate by using PH test paper, and the result shows that the PH is 6.5-7.0.

E. 72.77g Fmoc-Aib-OH, 33.93g DIC and 36.33g HOBT were weighed in sequence in a clean 1L beaker, 300ml of DMF/DCM solution in a volume ratio of 1:1 was added, and the mixture was placed in ice water and stirred with a mechanical stirrer at 0-10℃for dissolution, and after the temperature was constant, the temperature was maintained and stirring was continued for 5min. Slowly adding the activating solution into a reaction kettle, and mixing and reacting for 2 hours at 20-25 ℃. After the reaction was completed, the mixture was drained, 300ml of DMF was added thereto, and after mixing for 5 minutes, the mixture was drained. The DMF washes were repeated 5 times, 300ml each time, and 5min each time of mixing. Finally, detecting the obtained product as negative by ninhydrin to obtain Fmoc-Aib-Glu (OBzl) -CTC resin.

F. The amino acid Fmoc-His (Boc) -OH was coupled as described above for the deprotection method of step D and the coupling method of step E. Finally, washing with dichloromethane for 5 times, 300ml each time; after washing, the mixture is washed twice with 300ml of methanol each time; washing with dichloromethane for 2 times, 300ml each time; finally, the resin was washed 3 times with 300ml portions of methanol until the resin was sufficiently dispersed. The resin was dried in a vacuum oven at 20-30 ℃ for 4h to constant weight (twice weighing in succession, error lower than 1%). 185.76g of a fully-protected Fmoc-His (Boc) -Aib-Glu (OBzl) -CTC resin was obtained.

G. The lysate ratio is TFE: DCM=1:4 (volume ratio), 30g of the full protection peptide resin of the CTC resin obtained in the step F is added into 300mL of lysate at 15 ℃, the temperature is raised to 30 ℃, the reaction is continued for 3 hours by stirring, then the filtered resin is filtered by a sand core funnel, the filtered resin is washed by 150mL of DCM, the filtrate is combined after repeated operation for two times, the filtrate is concentrated under reduced pressure until the volume of the filtrate is 30% of the original volume, then the concentrated solution is slowly added into precooled 1L of isobutyl ether, the mixture is centrifuged for 5 times after sedimentation, 200mL of isobutyl ether is used each time, white solid powder is obtained, the mixture is dried by nitrogen gas firstly, then dried by a vacuum drying box for 10 hours, and 12.95g of full protection Fmoc-His (Boc) -Aib-Glu (OBzl) -OH crude product is obtained after weighing.

EXAMPLE 3 preparation of Fmoc-His (Trt) -Aib-Glu (OtBu) -OH

A. 120g of 2-CTC resin with substitution degree of 1.06mmol/g is added into a reaction kettle, 400ml of dichloromethane is added, after mixing for 2min, the dichloromethane is filtered off, 400ml of dichloromethane is added again, after mixing for 40min, the dichloromethane is filtered off, finally 400ml of dichloromethane is added again, after mixing for 2min, the dichloromethane is filtered off, and the resin is ready for use.

B. 108.24g of Fmoc-Glu (OtBu) -OH was weighed into a beaker, 400ml of DMF and 63.06ml of DIEA were added, the solution was stirred at 0-10℃for 5min and then poured into the CTC resin obtained in step A, and the mixture was reacted for 4h at 20-25 ℃. After the reaction is finished, DMF is filtered off. A mixed solution of 20ml of methanol and 200ml of DMF, a mixed solution of 30ml of DIEA and 200ml of DMF was added to the resin, and the mixing reaction was continued for 1h. After the reaction is finished, carrying out suction filtration, and washing the resin with DMF for 5 times, wherein 400ml of the resin is used each time; after washing, methanol is used for washing twice, 400ml each time; washing with dichloromethane for 2 times, 400ml each time; finally, the resin was washed 3 times with 400ml portions of methanol until the resin was sufficiently dispersed.

C. And (3) drying the resin obtained in the step (B) in a vacuum drying oven at 20-30 ℃ for 4 hours until the weight is constant (twice weighing is carried out continuously, and the error is lower than 1%). After drying, 175.21g of Fmoc-Glu (OtBu) -CTC resin was obtained, and the degree of substitution was detected to be 0.73mmol/g.

D. The Fmoc-Glu (OtBu) -CTC resin obtained in step C was poured into a reaction vessel, swollen with 400ml DCM, mixed for 15min and then pumped down. 500ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 5min, followed by draining. DMF400ml was added and after mixing for 5min, the mixture was drained. 400ml of 20% piperidine/DMF solution was added and mixed at 20-30℃for 10min, followed by draining. DMF400ml was added and after mixing for 5min, the mixture was drained. Repeated DMF washing for 5 times, 400ml each time, mixing for 5min each time, and after the 4 th washing, detecting the filtrate by using PH test paper, and the result shows that the PH is 6.5-7.0.

E. 82.71g Fmoc-Aib-OH, 38.53g DIC and 41.26g HOBT were weighed in sequence in a clean 1L beaker, 400ml of DMF/DCM solution in a volume ratio of 1:1 was added, and the mixture was placed in ice water and stirred with a mechanical stirrer at 0-10℃for dissolution, and after the temperature was constant, the temperature was maintained and stirring was continued for 5min. Slowly adding the activating solution into a reaction kettle, and mixing and reacting for 2 hours at 20-25 ℃. After the reaction was completed, the mixture was drained, and after adding 400ml of DMF and mixing for 5 minutes, the mixture was drained. The DMF washes were repeated 5 times, 400ml each time, and 5min each time of mixing. Finally, detecting the obtained product as negative by ninhydrin to obtain Fmoc-Aib-Glu (OtBu) -CTC resin.

F. The amino acid Fmoc-His (Trt) -OH was coupled as described above for the deprotection method of step D and the coupling method of step E. Finally, washing with dichloromethane for 5 times, each time 400ml; after washing, methanol is used for washing twice, 400ml each time; washing with dichloromethane for 2 times, 400ml each time; finally, the resin was washed 3 times with 400ml portions of methanol until the resin was sufficiently dispersed. The resin was dried in a vacuum oven at 20-30 ℃ for 4h to constant weight (twice weighing in succession, error lower than 1%). 208.64g of a fully-protected Fmoc-His (Trt) -Aib-Glu (OtBu) -CTC resin was obtained.

G. The lysate ratio is TFE: DCM=1:4 (volume ratio), 20g of the full-protection peptide resin of the CTC resin obtained in the step F is added into 200mL of lysate at 15 ℃, the temperature is raised to 30 ℃, the reaction is continued for 3 hours by stirring, then the filtered resin is filtered by a sand core funnel, the filtered resin is washed by 100mL of DCM, the filtrate is combined after repeated operation for two times, the filtrate is concentrated under reduced pressure until the volume of the filtrate is 30% of the original volume, then the concentrated solution is slowly added into precooled 1L of isobutyl ether, the mixture is centrifuged for 5 times after sedimentation, 200mL of isobutyl ether is used each time, white solid powder is obtained, the mixture is dried by nitrogen gas firstly, then dried by a vacuum drying box for 10 hours, and 8.25g of the full-protection Fmoc-His (Trt) -Aib-Glu (OtBu) -OH crude product is obtained after weighing.

EXAMPLE 4 preparation of Fmoc-Gly-Thr (tBu) -Phe-OH

A. 150g of 2-CTC resin with substitution degree of 1.10mmol/g is added into a reaction kettle, 500ml of dichloromethane is added, after mixing for 2min, dichloromethane is filtered off, 500ml of dichloromethane is added again, after mixing for 40min, dichloromethane is filtered off, finally 500ml of dichloromethane is added again, after mixing for 2min, dichloromethane is filtered off, and the resin is ready for use.

B. 127.85g of Fmoc-Phe-OH was weighed into a beaker, 500ml of DMF and 81.81ml of DIEA were added, the solution was stirred and activated at 0-10℃for 5min, then poured into the CTC resin obtained in step A, and mixed and reacted at 20-25℃for 4h. After the reaction is finished, DMF is filtered off. A mixed solution of 25ml of methanol and 250ml of DMF, a mixed solution of 40ml of DIEA and 250ml of DMF was added to the resin, and the mixing reaction was continued for 1h. After the reaction is finished, carrying out suction filtration, and washing the resin with DMF for 5 times, wherein 500ml of the resin is used each time; after washing, methanol is used for washing twice, and 500ml of methanol is used for each time; washing with dichloromethane for 2 times, each time with 500ml; finally, the resin was washed 3 times with 500ml portions of methanol until the resin was sufficiently dispersed.

C. And (3) drying the resin obtained in the step (B) in a vacuum drying oven at 20-30 ℃ for 4 hours until the weight is constant (twice weighing is carried out continuously, and the error is lower than 1%). After drying, 205.46g of Fmoc-Phe-CTC resin were obtained, the degree of substitution detected being 0.80mmol/g.

D. The Fmoc-Phe-CTC resin obtained in step C was poured into the reaction vessel, swollen with 500ml DCM and mixed for 15min, and then pumped down. 500ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 5min, followed by draining. DMF500ml was added and after mixing for 5min, the mixture was drained. 500ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 10min, followed by draining. DMF500ml was added and after mixing for 5min, the mixture was drained. Repeated DMF washing 5 times, 500ml each time, mixing 5min each time, and after washing 4 times, the filtrate was checked with PH test paper, which showed that PH was acceptable at 6.5-7.0.

E. 131.16g Fmoc-Thr (tBu) -OH, 49.98g DIC and 53.51g HOBT were weighed in sequence in a clean 1L beaker, 500ml of DMF/DCM solution was added in a volume ratio of 1:1, and placed in ice water and stirred with a mechanical stirrer at 0-10℃for dissolution, after the temperature was constant, the temperature was maintained and stirring was continued for 5min. Slowly adding the activating solution into a reaction kettle, and mixing and reacting for 2 hours at 20-25 ℃. After the reaction, the mixture was drained, 500ml of DMF was added thereto, and after 5 minutes of mixing, the mixture was drained. The DMF washes were repeated 5 times, 500ml each time, and 5min each time of mixing. Finally, detecting the obtained product as negative by ninhydrin to obtain Fmoc-Thr (tBu) -Phe-CTC resin.

F. The amino acid Fmoc-Gly-OH was coupled as described above for the deprotection method of step D and the coupling method of step E. Finally, washing with dichloromethane for 5 times, each time 500ml; after washing, methanol is used for washing twice, and 500ml of methanol is used for each time; washing with dichloromethane for 2 times, each time with 500ml; finally, the resin was washed 3 times with 500ml portions of methanol until the resin was sufficiently dispersed. The resin was dried in a vacuum oven at 20-30 ℃ for 4h to constant weight (twice weighing in succession, error lower than 1%). 236.15g of a fully protected Fmoc-Gly-Thr (tBu) -Phe-CTC resin was obtained.

G. The mixture ratio of the lysate is TFE (TFE) =1:4 (volume ratio), 20g of the full-protection peptide resin of the CTC resin obtained in the step F is added into 200mL of lysate at 15 ℃, the temperature is raised to 30 ℃, the reaction is continued for 3 hours by stirring, then the filtered resin is filtered by a sand core funnel, the filtered resin is washed by 100mL of DCM, the filtrate is combined after repeated operation for two times, the filtrate is concentrated under reduced pressure until the volume of the filtrate is 30% of the original volume, then the concentrated solution is slowly added into precooled 1L of isobutyl ether, the mixture is centrifuged for 5 times after sedimentation, 200mL of isobutyl ether is used each time, white solid powder is obtained, the mixture is dried by nitrogen, then dried by a vacuum drying box for 10 hours, and 7.39g of the full-protection Fmoc-Gly-Thr (tBu) -Phe-OH crude product is obtained after weighing.

EXAMPLE 5 Fmoc-Gly-Thr (Bzl) -Phe-OH preparation

A. 100g of 2-CTC resin with substitution degree of 1.10mmol/g is added into a reaction kettle, 300ml of dichloromethane is added, after mixing for 2min, dichloromethane is filtered off, 300ml of dichloromethane is added again, after mixing for 40min, dichloromethane is filtered off, finally 300ml of dichloromethane is added again, after mixing for 2min, dichloromethane is filtered off, and the resin is ready for use.

B. 85.23g Fmoc-Phe-OH was weighed into a beaker, 300ml DMF and 54.54ml DIEA were added, the solution was stirred and activated at 0-10℃for 5min, then poured into the CTC resin obtained in step A, and mixed and reacted at 20-25℃for 4h. After the reaction is finished, DMF is filtered off. A mixed solution of 20ml of methanol and 150ml of DMF, a mixed solution of 25ml of DIEA and 150ml of DMF was added to the resin, and the mixing reaction was continued for 1h. After the reaction is finished, carrying out suction filtration, and washing the resin with DMF for 5 times, wherein 300ml of the resin is used each time; after washing, the mixture is washed twice with 300ml of methanol each time; washing with dichloromethane for 2 times, 300ml each time; finally, the resin was washed 3 times with 300ml portions of methanol until the resin was sufficiently dispersed.

C. And (3) drying the resin obtained in the step (B) in a vacuum drying oven at 20-30 ℃ for 4 hours until the weight is constant (twice weighing is carried out continuously, and the error is lower than 1%). After drying, 136.84g of Fmoc-Phe-CTC resin was obtained, the degree of substitution detected being 0.80mmol/g.

D. The Fmoc-Phe-CTC resin obtained in step C was poured into the reaction vessel, swollen with 300ml DCM and mixed for 15min, and then pumped down. 300ml of 20% piperidine/DMF solution was added and mixed at 20-30℃for 5min, followed by draining. DMF (300 ml) was added thereto, and after mixing for 5 minutes, the mixture was drained. 300ml of 20% piperidine/DMF solution was added and mixed at 20-30℃for 10min, followed by draining. DMF (300 ml) was added thereto, and after mixing for 5 minutes, the mixture was drained. Repeated DMF washing for 5 times, 300ml each time, mixing for 5min each time, and after the 4 th washing, detecting the filtrate by using PH test paper, and the result shows that the PH is 6.5-7.0.

E. 87.44g Fmoc-Thr (Bzl) -OH, 33.32g DIC and 35.67g HOBT were weighed in sequence in a clean 1L beaker, 300ml of DMF/DCM solution in a volume ratio of 1:1 was added, and the mixture was placed in ice water and stirred with a mechanical stirrer at 0-10℃for dissolution, and after the temperature was constant, the temperature was maintained and stirring was continued for 5min. Slowly adding the activating solution into a reaction kettle, and mixing and reacting for 2 hours at 20-25 ℃. After the reaction was completed, the mixture was drained, 300ml of DMF was added thereto, and after mixing for 5 minutes, the mixture was drained. The DMF washes were repeated 5 times, 300ml each time, and 5min each time of mixing. Finally, detecting the obtained product as negative by ninhydrin to obtain Fmoc-Thr (Bzl) -Phe-CTC resin.

F. The amino acid Fmoc-Gly-OH was coupled as described above for the deprotection method of step D and the coupling method of step E. Finally, washing with dichloromethane for 5 times, 300ml each time; after washing, the mixture is washed twice with 300ml of methanol each time; washing with dichloromethane for 2 times, 300ml each time; finally, the resin was washed 3 times with 300ml portions of methanol until the resin was sufficiently dispersed. The resin was dried in a vacuum oven at 20-30 ℃ for 4h to constant weight (twice weighing in succession, error lower than 1%). 160.86g of a fully protected Fmoc-Gly-Thr (Bzl) -Phe-CTC resin was obtained.

G. The mixture ratio of the lysate is TFE (TFE) =1:4 (volume ratio), 20g of the full-protection peptide resin of the CTC resin obtained in the step F is added into 200mL of lysate at 15 ℃, the temperature is raised to 30 ℃, the reaction is continued for 3 hours by stirring, then the filtered resin is filtered by a sand core funnel, the filtered resin is washed by 100mL of DCM, the filtrate is combined after repeated operation for two times, the filtrate is concentrated under reduced pressure until the volume of the filtrate is 30% of the original volume, then the concentrated solution is slowly added into precooled 1L of isobutyl ether, the mixture is centrifuged for 5 times after sedimentation, 200mL of isobutyl ether is used each time, white solid powder is obtained, the mixture is dried by nitrogen firstly, then dried by a vacuum drying box for 10 hours, and 7.84g of full-protection Fmoc-Gly-Thr (Bzl) -Phe-OH crude product is obtained after weighing.

EXAMPLE 6 preparation of Fmoc-Lys (AEEA-AEEA-gamma-Glu (OtBu) -Octadecananic) -OH

A. 150g of 2-CTC resin with substitution degree of 1.05mmol/g is added into a reaction kettle, 500ml of dichloromethane is added, after mixing for 2min, dichloromethane is filtered off, 500ml of dichloromethane is added again, after mixing for 40min, dichloromethane is filtered off, finally 500ml of dichloromethane is added again, after mixing for 2min, dichloromethane is filtered off, and the resin is ready for use.

B. 121.44g of Fmoc-AEEA-OH was weighed into a beaker, 500ml of DMF and 76.18ml of DIEA were added, the solution was stirred at 0-10℃for 5min, and then poured into the CTC resin obtained in step A, and mixed and reacted at 20-25℃for 4h. After the reaction is finished, DMF is filtered off. A mixed solution of 25ml of methanol and 250ml of DMF, a mixed solution of 40ml of DIEA and 250ml of DMF was added to the resin, and the mixing reaction was continued for 1h. After the reaction is finished, carrying out suction filtration, and washing the resin with DMF for 5 times, wherein 500ml of the resin is used each time; after washing, methanol is used for washing twice, and 500ml of methanol is used for each time; washing with dichloromethane for 2 times, each time with 500ml; finally, the resin was washed 3 times with 500ml portions of methanol until the resin was sufficiently dispersed.

C. And (3) drying the resin obtained in the step (B) in a vacuum drying oven at 20-30 ℃ for 4 hours until the weight is constant (twice weighing is carried out continuously, and the error is lower than 1%). After drying, 140.20 mmole Fmoc-AEEA-CTC resin was obtained, and the degree of substitution was detected to be 0.73 mmole/g.

D. The Fmoc-AEEA-CTC resin obtained in step C was poured into the reactor, swelled and mixed with 500ml DCM for 15min, and then pumped down. 500ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 5min, followed by draining. DMF500ml was added and after mixing for 5min, the mixture was drained. 500ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 10min, followed by draining. DMF500ml was added and after mixing for 5min, the mixture was drained. Repeated DMF washing 5 times, 500ml each time, mixing 5min each time, and after washing 4 times, the filtrate was checked with PH test paper, which showed that PH was acceptable at 6.5-7.0.

E. 108.07g Fmoc-AEEA-OH, 35.42g DIC and 39.59g HOBT were weighed in sequence in a clean 1L beaker, 500ml of DMF/DCM solution in a volume ratio of 1:1 was added, the mixture was placed in ice water and stirred with a mechanical stirrer at 0-10℃for dissolution, and after the temperature was constant, the temperature was maintained and stirring was continued for 5min. Slowly adding the activating solution into a reaction kettle, and mixing and reacting for 2 hours at 20-25 ℃. After the reaction, the mixture was drained, 500ml of DMF was added thereto, and after 5 minutes of mixing, the mixture was drained. The DMF washes were repeated 5 times, 500ml each time, and 5min each time of mixing. Finally, detecting the obtained product as negative by ninhydrin to obtain Fmoc-AEEA-AEEA-CTC resin.

F. According to the deprotection method of the step D and the coupling method of the step E, the amino acid Fmoc-Glu (OH) -OtBu and the octadecanedioic acid mono-tert-butyl ester are respectively coupled in sequence. Finally, washing with dichloromethane for 5 times, each time 500ml; after washing, methanol is used for washing twice, and 500ml of methanol is used for each time; washing with dichloromethane for 2 times, each time with 500ml; finally, the alcohol was used 3 times, 500ml each, until the resin was sufficiently dispersed. The resin was dried in a vacuum oven at 20-30 ℃ for 4h to constant weight (twice weighing in succession, error lower than 1%). 255.40g of Octadecanadioic-gamma-Glu (OtBu) -AEEA-AEEA-CTC resin was obtained.

G. The lysate ratio is TFE: DCM=1:4 (volume ratio), 30g of the full-protection peptide resin of the CTC resin obtained in the step F is added into 300mL of lysate at 15 ℃, the temperature is raised to 30 ℃, the reaction is continued for 3 hours by stirring, then the filtered resin is filtered by a sand core funnel, the filtered resin is washed by 100mL of DCM, the filtrate is combined after repeated operation for two times, the filtrate is concentrated under reduced pressure until the volume of the filtrate is 30% of the original volume, then the concentrated solution is slowly added into pre-cooled 2L of isobutyl ether, the mixture is centrifuged for 5 times after sedimentation, each time 300mL of isobutyl ether is used, white solid powder is obtained, the mixture is dried by nitrogen, then dried by a vacuum drying box for 10 hours, and 11.65g of the crude product of Octadecananic-gamma-Glu (OtBu) -EA-AEEA-OH is obtained after weighing.

H. 5G of crude Octadecanadic-gamma-Glu (OtBu) -AEEA-AeEA-OH obtained in step G was dissolved in 10mL of CM, and 2.2G of pentafluorophenol was added. 2.4g of DCC was weighed and dissolved in 10mM DCM, the DCC solution was slowly added dropwise to the reaction solution, the reaction was stirred for 1.0h, and after completion of the TLC detection, the reaction was filtered. The filtrate was washed once with saturated brine, once with water, and the DCM solution was dried over anhydrous sodium sulfate, concentrated to dryness, and dissolved in an appropriate amount of acetonitrile. 6.08g Fmoc-Lys-OH.HCl was weighed out and dissolved in acetonitrile/water (acetonitrile/water=1/2), 7.5ml of LDIEA was added and stirred for 15 minutes. The above reaction solution was slowly added dropwise to the Fmoc-Lys-OH solution and the reaction was stirred for 1.5h. Dilute hydrochloric acid was added to adjust the pH to about 6, and a small amount of DCM was added for extraction. Fmoc-Lys (AEEA-AEEA-. Gamma. -Glu (OtBu) -Octadecananic) -OH2.85g was obtained by purification.

EXAMPLE 7 preparation of Fmoc-Gly-Wang resin with substitution of 0.30mmol/g

A. 10g of Wang resin with a substitution degree of 0.90mmol/g was added to the reaction vessel, 100ml of dichloromethane was added, after mixing for 2min, dichloromethane was filtered off, 100ml of dichloromethane was added again, after mixing for 40min, dichloromethane was filtered off, finally 100ml of dichloromethane was added again, after mixing for 2min, dichloromethane was filtered off and the resin was ready for use.

B. 5.35g Fmoc-Gly-OH and 2.92g HOBT were weighed into a beaker, 100ml DMF and 4.46ml DIEA were added, the solution was stirred at 0-10℃for 5min, then poured into the Wang resin obtained in step A, 0.16g DMAP was added, and the mixture was mixed at 20-25℃for 4h. After the reaction was completed, 8.5ml of acetic anhydride was added thereto, and mixing was continued for 1 hour. After the reaction is finished, carrying out suction filtration, and washing the resin with DMF for 5 times, wherein 100ml of the resin is used each time; after washing, methanol is used for washing twice, and 100ml of methanol is used for each time; washing with dichloromethane for 2 times, each time with 100ml; finally, the resin was washed 3 times with 100ml portions of methanol until the resin was sufficiently dispersed.

C. And (3) drying the resin obtained in the step (B) in a vacuum drying oven at 20-30 ℃ for 4 hours until the weight is constant (twice weighing is carried out continuously, and the error is lower than 1%). After drying, 12.87g of Fmoc-Gly-Wang resin was obtained, and the substitution degree by ultraviolet detection was 0.30mmol/g.

EXAMPLE 8 preparation of crude Sodamantane peptide 1

A. 10g of Fmoc-Gly-Wang resin obtained in example 7 was poured into a reaction kettle, swollen with 100ml of DCM and mixed for 15min, and then pumped down. 100ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 5min, followed by draining. DMF (100 ml) was added thereto, and after mixing for 5 minutes, the mixture was drained. 100ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 10min, followed by draining. DMF (100 ml) was added thereto, and after mixing for 5 minutes, the mixture was drained. Washing with DMF was repeated 8 times, 100ml each time, mixing 5min each time, and after the seventh washing, the filtrate was checked with pH paper, which showed that the pH was acceptable at 6.5-7.0.

B. 3.89g of Fmoc-Arg (Pbf) -OH, 2.31g of TBTU and 0.97g of HOBT are weighed in sequence into a clean 1L beaker, 100ml of DMF/DCM solution with the volume ratio of 1:1 is added, the mixture is placed into ice water and stirred and dissolved by a mechanical stirrer at the temperature of 0-10 ℃, after the temperature is constant, 1.49ml of LDIEA is added, the temperature is continuously maintained and stirring and activation are carried out for 5min. Slowly adding the activating solution into a reaction kettle, and mixing for 2 hours at 20-25 ℃. After the reaction, the mixture was drained, 100ml of DMF was added thereto, and after 5 minutes of mixing, the mixture was drained. The DMF washes were repeated 6 times, 100ml each time, and 5min each time mixed. Finally, detecting the obtained product as negative by ninhydrin to obtain the Fmoc-Arg-Gly-Wang resin.

C. According to the deprotection method of the step A and the coupling method of the step B, the residual amino acids or peptide fragments are respectively coupled in sequence according to the sequence of main chain amino acids, namely: fmoc-Gly-OH, fmoc-Arg (Pbf) -OH, fmoc-Val-OH, fmoc-Leu-OH, fmoc-Trp (Boc) -OH, fmoc-Ala-OH, fmoc-Ile-OH, fmoc-Phe-OH, fmoc-Glu (OtBu) -OH, fmoc-Lys (AEEA-AEEA-gamma-Glu (OtBu) -Octadeceionic) -OH obtained in example 7, fmoc-Ala-OH, fmoc-Gln (Trt) -OH, fmoc-Gly-OH, fmoc-Glu (OtBu) -OH, fmoc-Leu-OH, fmoc-Tyr (tBu) -OH, fmoc-Ser (OtBu) -OH, fmoc-Asp (OtBu) -OH, fmoc-Asp-OH (Fmoc-OH) obtained in example 7, and Fmoc-Val-OH (Fmoc-OH) obtained in example 4. Thr-Boc-OH. After the coupling is completed, the reaction mixture is washed with dichloromethane for 5 times, 100ml each time; after washing, methanol is used for washing twice, and 100ml of methanol is used for each time; washing with dichloromethane for 2 times, each time with 100ml; finally, the resin was washed 3 times with 100ml of alcohol until the resin was sufficiently dispersed. The resin was dried in a vacuum oven at 20-30 ℃ for 4h to constant weight (twice weighing in succession, error lower than 1%). 26.15g of Somam Lu Taitai resin are obtained. HPLC chromatograms are shown in FIG. 1.

EXAMPLE 9 preparation of crude Sodamantane peptide 2

A. 10g of Fmoc-Gly-Wang resin with substitution of 0.30mmol/g was poured into the reaction kettle, swollen with 100ml of DCM and mixed for 15min, and then pumped down. 100ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 5min, followed by draining. DMF (100 ml) was added thereto, and after mixing for 5 minutes, the mixture was drained. 100ml of a 20% piperidine/DMF solution was added and mixed at 20-30℃for 10min, followed by draining. DMF (100 ml) was added thereto, and after mixing for 5 minutes, the mixture was drained. Washing with DMF was repeated 8 times, 100ml each time, mixing 5min each time, and after the seventh washing, the filtrate was checked with pH paper, which showed that the pH was acceptable at 6.5-7.0.

C. According to the deprotection method of the step A and the coupling method of the step B, the residual amino acids or peptide fragments are respectively coupled in sequence according to the sequence of main chain amino acids, namely: fmoc-Gly-OH, fmoc-Arg (Pbf) -OH, fmoc-Val-OH, fmoc-Leu-OH, fmoc-Trp (Boc) -OH, fmoc-Ala-OH, fmoc-Ile-OH, fmoc-Phe-OH, fmoc-Glu (OtBu) -OH, fmoc-Lys (AEEA-AEEA-gamma-Glu (OtBu) -Octadeceionic) -OH obtained in example 7, fmoc-Ala-OH, fmoc-Gln (Trt) -OH, fmoc-Gly-OH, fmoc-Glu (OtBu) -OH, fmoc-Leu-OH, fmoc-Tyr (tBu) -OH, fmoc-Ser (OtBu) -OH, fmoc-Asp (OtBu) -OH, fmoc-Asp-OH (Fmoc-OH) obtained in example 7, and Fmoc-Val-OH (Fmoc-OH) obtained in example 2. BtBu-OH. After the coupling is completed, the reaction mixture is washed with dichloromethane for 5 times, 100ml each time; after washing, methanol is used for washing twice, and 100ml of methanol is used for each time; washing with dichloromethane for 2 times, each time with 100ml; finally, the resin was washed 3 times with 100ml of alcohol until the resin was sufficiently dispersed. The resin was dried in a vacuum oven at 20-30 ℃ for 4h to constant weight (twice weighing in succession, error lower than 1%). 27.08g of Somam Lu Taitai resin are obtained. The HPLC chromatogram is similar to fig. 1.

EXAMPLE 10 preparation of crude Sodamantane peptide 3

C. According to the deprotection method of the step A and the coupling method of the step B, the residual amino acids or peptide fragments are respectively coupled in sequence according to the sequence of main chain amino acids, namely: fmoc-Gly-OH, fmoc-Arg (Pbf) -OH, fmoc-Val-OH, fmoc-Leu-OH, fmoc-Trp (Boc) -OH, fmoc-Ala-OH, fmoc-Ile-OH, fmoc-Phe-OH, fmoc-Glu (OtBu) -OH, fmoc-Lys (AEEA-AEEA-gamma-Glu (OtBu) -Octadeceioic) -OH, fmoc-Ala-OH, fmoc-Gln (Trt) -OH, fmoc-Gly-OH, fmoc-Glu (OtBu) -OH obtained in example 7 coupling of Fmoc-Glu (OtBu) -OH, fmoc-Leu-OH, fmoc-Tyr (tBu) -OH, fmoc-Ser (tBu) -OH, fmoc-Val-OH, fmoc-Asp (OtBu) -OH, fmoc-Ser (tBu) -OH, fmoc-Thr (tBu) -OH, fmoc-Gly-Thr (tBu) -Phe-OH obtained in example 4, and Fmoc-His (Trt) -Aib-Glu (OtBu) -OH obtained in example 3. After the coupling is completed, the reaction mixture is washed with dichloromethane for 5 times, 100ml each time; after washing, methanol is used for washing twice, and 100ml of methanol is used for each time; washing with dichloromethane for 2 times, each time with 100ml; finally, the resin was washed 3 times with 100ml of alcohol until the resin was sufficiently dispersed. The resin was dried in a vacuum oven at 20-30 ℃ for 4h to constant weight (twice weighing in succession, error lower than 1%). 26.91g of Somam Lu Taitai resin are obtained. The HPLC chromatogram is similar to fig. 1.

EXAMPLE 11 preparation of Sodamantane spermidine

The crude product of the somalundum obtained in the example 8 is dissolved in dilute ammonia water, the pH value of the crude solution of the somalundum is regulated to 8.0-8.5 by phosphoric acid, and the solution of the somalundum is obtained by filtering. Performing HPLC linear gradient elution by taking octalkyl bonded silica gel as a stationary phase and ammonium chloride and acetonitrile as flowing relative to the crude peptide solution of the somalundum, collecting a somalundum fraction, and removing part of acetonitrile by rotary evaporation by using a rotary evaporator to obtain a primary purified solution of the somalundum. The primary purification solution of the somalundum takes octaalkyl bonding silica gel as a stationary phase, potassium dihydrogen phosphate aqueous solution of which the pH is regulated by phosphoric acid, acetonitrile and isopropanol mixed solvent as a mobile phase are subjected to HPLC linear elution, a somalundum fraction is collected, a rotary evaporator is used for rotary evaporation to remove part of acetonitrile, and the secondary purification solution of the somalundum is obtained. The secondary purification liquid of the somalundum takes octaalkyl bonding silica gel as a stationary phase, ammonium bicarbonate aqueous solution and acetonitrile as a mobile phase to carry out HPLC linear elution, the somalundum fraction is collected, the acetonitrile and most of water are removed by rotary evaporation through rotary evaporator, 4.86g of the somalundum peptide refined peptide is obtained by freeze drying, the HPLC purity is 99.75%, the HPLC chromatogram is shown in figure 2, and the purification yield is 68.24%.

Claims

1. A solid phase synthesis method of a somalupeptide is characterized in that: respectively synthesizing R1-His (R2) -Aib-Glu (OR 3) -OH fragment and R4-Gly-Thr (R5) -Phe-OH fragment, using for synthesizing the somalupeptide resin, and obtaining the somalupeptide through cleavage and purification, wherein

R1 is selected from the group consisting of Fmoc, dde, alloc, boc, trt, dmb, mmt, mtt,

r2 is selected from the group consisting of Fmoc, dde, alloc, boc, trt, dmb, mmt, mtt,

r3 is selected from tBu and Bzl,

r4 is selected from Fmoc,

r5 is selected from tBu and Bzl.

2. The solid phase synthesis method of somalupeptide of claim 1, wherein: the R1 is selected from the group consisting of: boc or Fmoc, R2 is selected from: trt or Boc, R3 is selected from tBu, R4 is selected from Fmoc, and R5 is selected from tBu.

3. The solid phase synthesis method of somalupeptide of claim 2, wherein: r1 is Boc, R2 is Trt, R3 is tBu, R4 is Fmoc, and R5 is tBu.

4. The method of claim 1, wherein in the solid phase synthesis of somalundin, S20 is: fmoc-Lys (AEEA-AEEA-gamma-Glu (OtBu) -Octadecananioic) -OH.