A salt-changing method for telpotide

Abstract

The invention provides a salt-exchanging method of tepote, which adopts high performance liquid chromatography and comprises the following steps: S1) using mobile phase A1 and mobile phase B to reversely flush a balanced preparation column; S2) loading a sample to be salt-exchanged onto the preparation column in reverse; S3) using mobile phase A1 and mobile phase B to forwardly flush the preparation column; S4) using mobile phase A2 and mobile phase B to forwardly flush the preparation column; S5) using mobile phase A2 and mobile phase B to perform forward isocratic elution and collect samples; S6) concentrating and drying qualified samples in the collected samples in S5) to obtain salt-exchanged tepote; wherein mobile phase B is acetonitrile, ethanol or isopropanol; mobile phase A1 is a sodium salt solution with a pH value of 4.0 to 9.0; mobile phase A2 is water or acetic acid aqueous solution; and the internal filler in the preparation column is a reverse phase chromatography filler. The above method has a short production cycle, and the obtained tepote has high solubility, controllable pH and high concentration.

Description

Technical Field

[0001] The present invention relates to the field of polypeptide drug preparation and purification, and in particular to a salt-exchanging method for telpotide. Background Art

[0002] Tirzepatide, also known as Tirzepatide in English, is a macromolecular peptide drug. It is a peptide composed of 39 amino acids with a molecular formula of C 225 H 348 N 48 O 68 , with a molecular weight of 4813.45Da. Its peptide chain sequence is as follows: Tyr 1 -Aib-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Ile-Aib-Leu 14 -Asp-Lys-Ile-Ala-Gln-Lys[tBuO-Eic-γ-Glu(OtBu)-AEEA-AEEA]-Ala 21 -Phe-Val-Gln-Trp-Leu-Ile-Ala-Gly 29 -Gly-Pro-Ser-Ser-Gly-Ala-Pro-Pro-Pro-Ser 39 -NH 2 The peptide chain contains more Asp and Glu, is an acidic peptide, has an isoelectric point of 4.6, and is very easy to bind to cations.

[0003] Telpotide is an agonist that acts on both GIP and GLP-1 receptors. Its mechanism of action is similar to that of GLP-1 receptor agonists, but it has an additional GIP agonist effect, so it is called a dual agonist. Research data show that in patients with type 2 diabetes, telpotide improves beta cell function and insulin sensitivity markers, significantly reduces blood sugar levels, significantly reduces body weight, improves gastrointestinal side effects, and also improves non-alcoholic fatty liver disease (NASH) markers. The original research and various references all use liquid phase splicing synthesis to synthesize telpotide. After purification, the finished product is obtained by precipitation methods such as tangential membrane filtration and methyl tert-ether co-feed precipitation. The finished product obtained by this salt exchange method has problems such as low yield, low peptide content, large residue, and uneven properties.

[0004] Patent CN 110903355 A Shengnuo Biopharmaceutical Co., Ltd. has announced a salt exchange method for Tirzepatide. It includes the following steps: Step 1: Purification by high performance liquid chromatography, using 0.1% TFA / water solution-acetonitrile solution and 50mmol ammonium acetate / water solution-acetonitrile to purify the purified product in turn; Step 2: Salt exchange by high performance liquid chromatography, the mobile phase system is 1% acetic acid / water solution-acetonitrile; gradient elution and cyclic loading method are used; Step 3: Combine the main peak solution of salt exchange, concentrate under reduced pressure, obtain Tirzepatide acetic acid aqueous solution, freeze-dry, and obtain 37.9g ​​of pure Tirzepatide, with a total yield of 26.2% and a molecular weight of 4813.6.

[0005] Since the isoelectric point of the peptide chain is 4.6, it is easy to collect cations. The product obtained by salt exchange according to the above patent contains ammonium and acetate, and the fraction solution volume produced by gradient elution and cyclic loading is large and difficult to concentrate. It is easy to precipitate after concentration, and the final product after reduced pressure concentration has low solubility and acetic acid residue.

[0006] Patent CN 112661815 A Nuotai Aosino Biopharmaceutical Co., Ltd. has announced a salt exchange method for Tirzepatide. It includes the following steps: Step 1: Purification by high performance liquid chromatography, using 0.2% ammonium perchlorate / water solution-acetonitrile solution and 20mmol sodium chloride / water solution-acetonitrile to purify the purified product in turn; Step 2: Combine the main peak solution, concentrate under reduced pressure to 20mg / ml, obtain Tirzepatide solution, freeze-dry, and obtain 5.4g of pure Tirzepatide, with a purification yield of 66.7% and a molecular weight of 4813.6.

[0007] Tipoline is an acidic peptide. The product obtained by salt exchange according to the above patent is rich in sodium ions and chloride ions, and there is no separate salt exchange procedure, which inevitably leads to a large volume of the output fraction solution and difficulty in concentration. It is easy to dissolve after concentration, but the final product of reduced pressure concentration has low solubility and high pH, ​​which is easy to produce degradation impurities. Summary of the invention

[0008] In order to solve the problems existing in the prior art such as the difficulty in concentration, large residual methyl tertiary ether or acetic acid, high or uncontrollable pH leading to the generation of degradation impurities, the present invention provides a salt exchange method for tepote, which can ensure that the purity and content of the sample remain unchanged, and the obtained tepote solid has high solubility for re-dissolution, controllable pH and high concentration, short production cycle of the entire process, less waste liquid, and is beneficial to environmental protection.

[0009] One aspect of the present invention provides a method for salt exchange of telpotide, wherein the salt exchange method is carried out by high performance liquid chromatography, and comprises the following steps:

[0010] S1) flushing the equilibrated preparative column with mobile phase A1 and mobile phase B in reverse order;

[0011] S2) reversely loading the purified tepoxetine sample to be replaced with salt onto a preparative column;

[0012] S3) flushing the preparative column with mobile phase A1 and mobile phase B in a forward direction;

[0013] S4) flushing the preparative column in the forward direction with mobile phase A2 and mobile phase B;

[0014] S5) performing isocratic elution in the forward direction with mobile phase A2 and mobile phase B, and collecting samples;

[0015] S6) concentrating and drying the qualified samples among the samples collected in S5) to obtain salt-exchanged tepoxetine;

[0016] Wherein, the mobile phase B is acetonitrile, ethanol or isopropanol;

[0017] The mobile phase A1 is a sodium salt solution with a pH value of 4.0 to 9.0;

[0018] The mobile phase A2 is water or acetic acid aqueous solution, and the concentration of acetic acid is 0.01% to 3.0%;

[0019] The filler in the preparation column is a reverse phase chromatography filler.

[0020] Furthermore, the reverse phase chromatography filler is preferably any one of C18, C8 or C4.

[0021] Furthermore, the particle size of the filler in the preparation column is 5 μm to 10 μm, and the pore size of the filler is

[0022] Further, the sodium salt solution is selected from any one of sodium hydroxide, sodium acetate or sodium bicarbonate, or a buffered saline solution containing sodium salt; preferably, the buffered saline solution containing sodium salt is selected from a buffered saline solution containing sodium dihydrogen phosphate and disodium hydrogen phosphate, a buffered saline solution containing phosphoric acid and sodium hydroxide, a buffered saline solution containing acetic acid and sodium acetate, a buffered saline solution containing sodium bicarbonate and sodium carbonate, a buffered saline solution containing acetic acid and sodium bicarbonate, a buffered saline solution containing bicarbonate and sodium hydroxide, and a buffered saline solution containing citric acid and sodium citrate.

[0023] Furthermore, the concentration of the mobile phase A1 sodium salt solution is 5 mmol / L to 500 mmol / L, preferably, the sodium salt concentration is 20 mmol / L to 200 mmol / L, for example, 50 mmol / L, 100 mmol / L, 150 mmol / L.

[0024] Furthermore, the pH value of the mobile phase A1 sodium salt solution is 4.0-9.0 without adjusting the pH value or adjusting the pH value to 4.0-9.0 with acetic acid, ammonia water, hydrochloric acid or potassium hydroxide, for example, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5.

[0025] Furthermore, the concentration of acetic acid in the mobile phase A2 is 0.02% to 1%, for example, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, or 0.9%.

[0026] Furthermore, in step S1), the mobile phase used for equilibrating the preparative column comprises 90%-95% of mobile phase A1 and 10%-5% of mobile phase B by volume.

[0027] Furthermore, in step S2), the mass of the sample is 1.0% to 10.0% of the mass of the reversed-phase chromatographic filler, preferably, the mass of the sample is 3.0% to 8.0% of the mass of the reversed-phase chromatographic filler, for example 4%, 5%, 6%, 7%, 8%.

[0028] Furthermore, in step S2), the reverse loading is to inject the telpotide sample to be replaced with salt from the outlet end of the chromatographic column.

[0029] Furthermore, in step S3), the mobile phase ratio used for flushing the preparation column is 90%-95% mobile phase A1 and 10%-5% mobile phase B by volume.

[0030] Furthermore, in step S4), the mobile phase ratio used for flushing the preparation column is 90%-95% mobile phase A2 and 10%-5% mobile phase B by volume.

[0031] Furthermore, in step S5), the mobile phase ratio used for eluting the preparation column is 30% to 60% mobile phase A2 and 70% to 40% mobile phase B by volume; further, the mobile phase ratio used is 50% mobile phase A2 and 50% mobile phase B by volume.

[0032] Furthermore, in step S6), the concentration and drying method is any one of rotary evaporation, thin film evaporation or spray drying.

[0033] Furthermore, in step S6), qualified samples are samples with a purity of ≥99.0% and a single impurity of ≤0.15%.

[0034] Another aspect of the present invention provides salt-exchanged telpotide prepared by the above-mentioned salt-exchange method.

[0035] Furthermore, the solubility of the salt-exchanged telpotide is above 100 mg / mL, and the pH after dissolution is 5-8.

[0036] Beneficial Effects

[0037] The present invention provides a salt-exchanging method for telpotide, which utilizes a special mobile phase combination, a sample loading method, and balancing, flushing, and elution conditions to achieve high solubility of telpotide reconstitution, controllable pH, high concentration, short production cycle of the entire process, and less waste liquid under the premise of ensuring that the purity and content of the sample remain unchanged. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 It is the HPLC chromatogram of the product of Example 1.

[0039] Figure 2 It is the HPLC chromatogram of the product of Example 2.

[0040] Figure 3 It is the HPLC chromatogram of the product of Example 3.

[0041] Figure 4 The chromatogram is the HPLC chromatogram of the product of Comparative Example 1.

[0042] Figure 5 It is the HPLC chromatogram of the product of Comparative Example 2.

[0043] Figure 6 The chromatogram is the HPLC chromatogram of the product of Comparative Example 3. DETAILED DESCRIPTION

[0044] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and understandable, the specific implementation modes of the present invention are described in detail below, but it should not be understood as limiting the applicable scope of the present invention.

[0045] In the present invention, "forward", "forward flushing" and the like refer to a method of using a conventional method of flushing, balancing or eluting a chromatographic column, injecting from the inlet end of the chromatographic column and flowing out from the outlet end of the chromatographic column.

[0046] In the present invention, "reverse", "reverse flushing" and the like refer to a method of injecting from the outlet of the chromatographic column and flowing out from the inlet of the chromatographic column. Reverse loading is a method of injecting the telpotide sample to be replaced with salt from the outlet of the chromatographic column.

[0047] In some specific embodiments of the present invention, a method for salt exchange of telpotide is provided, wherein the salt exchange is performed by high performance liquid chromatography.

[0048] In high performance liquid chromatography, the mobile phase B is selected from acetonitrile, ethanol or isopropanol.

[0049] Mobile phase A1 is selected from a sodium salt solution with a pH value of 4.0 to 9.0; the sodium salt solution is selected from any one of sodium hydroxide, sodium acetate or sodium bicarbonate, or a buffered salt solution containing sodium salt; the buffered salt solution containing sodium salt is selected from a buffered salt solution containing sodium dihydrogen phosphate and disodium hydrogen phosphate, a buffered salt solution containing phosphoric acid and sodium hydroxide, a buffered salt solution containing acetic acid and sodium acetate, a buffered salt solution containing sodium bicarbonate and sodium carbonate, a buffered salt solution containing acetic acid and sodium bicarbonate, a buffered salt solution containing bicarbonate and sodium hydroxide, and a buffered salt solution containing citric acid and sodium citrate. The concentration of the sodium salt solution in mobile phase A1 is 5 mmol / L to 500 mmol / L, and in a preferred embodiment, the sodium salt concentration is 20 mmol / L to 200 mmol / L. The pH value of the sodium salt solution of mobile phase A1 is 4.0 to 9.0 without adjusting the pH value or adjusting the pH to 4.0 to 9.0 with acetic acid, ammonia water, hydrochloric acid or potassium hydroxide.

[0050] The mobile phase A2 is selected from water or acetic acid aqueous solution, and the concentration of acetic acid is 0.01% to 3.0%. In a preferred embodiment, the concentration of acetic acid in the mobile phase A2 is 0.02% to 1%.

[0051] The packing in the preparation column is a reverse phase chromatography packing. In a preferred embodiment, the reverse phase chromatography packing is any one of C18, C8 or C4. The particle size of the packing in the preparation column is 5 μm to 10 μm, and the pore size of the packing is

[0052] The salt-exchanging method comprises the following steps:

[0053] S1) flushing the equilibrated preparative column with mobile phase A1 and mobile phase B in reverse order;

[0054] The mobile phase used for balancing the preparative column in step S1) is 90%-95% mobile phase A1 and 10%-5% mobile phase B by volume.

[0055] S2) Reverse loading the purified telpotide sample to be replaced with salt onto the preparation column; reverse loading is injecting the telpotide sample to be replaced with salt from the outlet end of the chromatographic column.

[0056] The purified telpotide sample to be replaced with salt is a telpotide sample purified by the same preparation column, and the mobile phase used in the purification process is acetonitrile as the organic phase, acetic acid and water as the aqueous phase; or acetonitrile as the organic phase, sodium acetate and acetic acid / ammonia water as the aqueous phase.

[0057] In step S2), the mass of the sample loaded is 1.0% to 10.0% of the mass of the reverse phase chromatography filler. In a preferred embodiment, the mass of the sample loaded is 3.0% to 8.0% of the mass of the reverse phase chromatography filler.

[0058] S3) flushing the preparative column with mobile phase A1 and mobile phase B in a forward direction;

[0059] The mobile phase used for flushing the preparation column in step S3) is composed of 90%-95% mobile phase A1 and 10%-5% mobile phase B by volume.

[0060] S4) flushing the preparative column in the forward direction with mobile phase A2 and mobile phase B;

[0061] The mobile phase used for flushing the preparation column in step S4) is 90%-95% mobile phase A2 and 10%-5% mobile phase B by volume.

[0062] S5) performing forward isocratic elution with mobile phase A2 and mobile phase B, forward flushing, and collecting samples;

[0063] In step S5), the elution direction is forward flushing, and the mobile phase ratio used is 30% to 60% mobile phase A2 and 70% to 40% mobile phase B by volume. In a preferred embodiment, 50% mobile phase A2 and 50% mobile phase B are used.

[0064] S6) Concentrating and drying the qualified samples among the samples collected in S5) to obtain salt-exchanged tepoxetine; the qualified samples are samples with a purity of ≥99.0% and a single impurity of ≤0.15%.

[0065] In step S6), the concentration and drying method is any one of rotary evaporation, thin film evaporation or spray drying.

[0066] Example 1

[0067] Weigh 200.0 g of crude telpotide (containing 121.1 g of telpotide), dissolve and filter, and purify through a reverse chromatographic preparative column to obtain a salt fraction product to be replaced, and concentrate the salt fraction product to be replaced to obtain a salt loading product to be replaced;

[0068] The reverse phase chromatography preparative column is 15 cm long and filled with 3.0 kg of C4 filler ( The average particle size is 10 μm and the average pore size is ), the purified telpotide sample to be replaced with salt is the telpotide sample purified by the same preparation column, and the mobile phase of the purification process uses acetonitrile as the organic phase, and acetic acid and water as the aqueous phase.

[0069] The salt is exchanged using an ultra-high performance liquid chromatograph and the same preparative column. The steps of the salt exchange include:

[0070] S1) Backwash the preparative column with 95% mobile phase A1 and 5% mobile phase B for 20 min at a flow rate of 500 mL / min; mobile phase A1 is a 40 mmol / L sodium acetate aqueous solution with acetic acid adjusting the pH to 8.0, and mobile phase B is isopropanol; backwashing means that the direction of mobile phase flow is opposite to that in normal use, that is, the mobile phase flows in from the outlet end of the preparative column and flows out from the inlet end.

[0071] S2) reversely loading the purified salt-exchange sample product onto the preparation column; reverse loading is injecting the salt-exchange sample product from the outlet end of the chromatographic column;

[0072] S3) flushing the preparative column with 95% mobile phase A1 and 5% mobile phase B in the forward direction for 40 min;

[0073] S4) flushing the preparative column in the forward direction with 95% mobile phase A2 and 5% mobile phase B for 40 min; A2 is sterile purified water;

[0074] S5) isocratically eluting with 50% mobile phase A2 and 50% mobile phase B in the forward direction to collect samples;

[0075] S6) Concentrating the qualified samples with purity ≥ 99.0% and single impurity ≤ 0.15% in the samples collected in S5) under reduced pressure, and freeze-drying to obtain 99.0 g of tepoxetine.

[0076] The salt conversion yield is 99.0%, the purification recovery rate is 81.8%, the purity is 99.25%, and the maximum single impurity is 0.11%. The freeze-dried product is dissolved, and the solubility reaches 120 mg / mL, and the pH is 7.61.

[0077] Example 2

[0078] Weigh 90.0 g of crude telpotide (containing 56.7 g of telpotide), dissolve and filter, and purify through a reverse chromatographic preparative column to obtain a salt fraction product to be replaced, and concentrate the salt fraction product to be replaced to obtain a salt loading product to be replaced;

[0079] The reverse phase chromatography preparative column is 10 cm and filled with 1.5 kg of C8 filler. The dynamic axial compression column (DAC) of the invention is used, the purified telpotide sample to be salt-exchanged is the telpotide sample purified by the same preparation column, and the mobile phase used in the purification process is acetonitrile as the organic phase, and sodium acetate and acetic acid / ammonia water as the aqueous phase.

[0080] The same preparation column is used for salt exchange, and the steps of salt exchange include:

[0081] S1) flushing the preparative column with 90% mobile phase A1 and 10% mobile phase B for 20 min at a flow rate of 200 mL / min; mobile phase A1 is a 40 mmol / L sodium bicarbonate aqueous solution adjusted to pH 8.0 with hydrochloric acid, and mobile phase B is acetonitrile;

[0082] S2) reversely loading the purified salt-exchange sample product onto the preparation column;

[0083] S3) flushing the preparative column with 95% mobile phase A1 and 5% mobile phase B in the forward direction for 40 min;

[0084] S4) flushing the preparative column in the forward direction with 90% mobile phase A2 and 10% mobile phase B for 20 min; A2 is 0.1% acetic acid aqueous solution;

[0085] S5) eluting in a forward isocratic manner with 40% mobile phase A2 and 60% mobile phase B, collecting the sample;

[0086] S6) Concentrating the qualified samples with purity ≥ 99.0% and single impurity ≤ 0.15% in the samples collected in S5) under reduced pressure, and freeze-drying to obtain 48.4 g of tepoxetine.

[0087] The conversion yield was 97.9%, the purification recovery was 85.4%, the purity was 99.30%, and the maximum single impurity was 0.12%. The freeze-dried product was dissolved, and the solubility reached 120 mg / mL, and the pH was 6.88.

[0088] Example 3-Comparative Example 4

[0089] The inventors used different conditions to carry out Examples 3-4 and Comparative Examples 1-3. The preparation columns were all 10 cm thick and filled with 1.5 kg of C8 filler. The differences between the A1 phase, the A2 phase and the sample loading method and those in Example 1 are summarized in the following table.

[0090]

[0091]

[0092]

[0093] By comparing the results of the embodiments and comparative examples, it can be seen that, firstly, a sodium salt aqueous solution is required in the mobile phase A1 used in the equilibrium preparation column. The finished product obtained after using the sodium salt aqueous solution has good solubility, while the product of comparative example 1 which does not use sodium salt will precipitate the sample after elution after reverse loading, and the solubility after freeze-drying is low, which is only 0.5 mg / mL.

[0094] However, only by setting the mobile phase A1 to contain sodium salt, there is a risk of high pH. For example, in Comparative Example 2, the pH value of the product after dissolution reaches 10.3, and the pH is high, which is very easy to produce degradation impurities. For this problem, the present invention adjusts the mobile phase to be rinsed with mobile phase A2, and controls the pH value of the A1 phase to control the pH and control the sodium ions. In addition, the solution concentration of the mobile phase A2 also needs to be controlled. If an acidic aqueous solution is used and the concentration is too high, the pH value of the final product will be too low. For example, in Comparative Example 3, it will also cause the problem of low solubility.

[0095] In addition, the forward washing and elution after reverse loading can shorten the cycle, control the sodium ions, and obtain a high-concentration, clear and transparent product. Non-reverse loading, that is, using the conventional forward loading method, has a low sample concentration, a longer production cycle, and obtains a single fixed sodium content product.

[0096] The present invention adopts the means of reverse loading and adjusting the mobile phase, and the two means complement each other to finally obtain a qualified freeze-dried sample with high solubility and adjustable pH after dissolution.

Claims

1. A salt-exchanging method for telportopeptide, wherein the salt-exchanging method is carried out by high performance liquid chromatography, characterized in that: It includes the following steps: S1) flushing the equilibrated preparative column with mobile phase A1 and mobile phase B in reverse order; S2) reversely loading the purified tepoxetine sample to be replaced with salt onto a preparative column; S3) flushing the preparative column with mobile phase A1 and mobile phase B in a forward direction; S4) flushing the preparative column with mobile phase A2 and mobile phase B in a forward direction; S5) performing isocratic elution in the forward direction with mobile phase A2 and mobile phase B, and collecting samples; S6) concentrating and drying the qualified samples among the samples collected in S5) to obtain salt-exchanged tepoxetine; Wherein, the mobile phase B is acetonitrile, ethanol or isopropanol; The mobile phase A1 is a sodium salt solution with a pH value of 4.0 to 9.0; The mobile phase A2 is water or acetic acid aqueous solution, and the concentration of acetic acid is 0.01% to 3.0%; The filler in the preparation column is a reverse phase chromatography filler.

2. The salt exchange method according to claim 1, characterized in that The sodium salt solution is selected from any one of sodium hydroxide, sodium acetate or sodium bicarbonate, or a buffered salt solution containing sodium salt; Preferably, the buffered saline solution containing sodium salt is selected from a buffered saline solution containing sodium dihydrogen phosphate and disodium hydrogen phosphate, a buffered saline solution containing phosphoric acid and sodium hydroxide, a buffered saline solution containing acetic acid and sodium acetate, a buffered saline solution containing sodium bicarbonate and sodium carbonate, a buffered saline solution containing acetic acid and sodium bicarbonate, a buffered saline solution containing bicarbonate and sodium hydroxide, and a buffered saline solution containing citric acid and sodium citrate.

3. The salt-exchanging method according to claim 1, characterized in that The concentration of the mobile phase A1 sodium salt solution is 5mmol / L to 500mmol / L; Preferably, the sodium salt concentration is 20 mmol / L to 200 mmol / L; Preferably, the pH value of the mobile phase A1 sodium salt solution is 4.0 to 9.0 without adjusting the pH value or adjusting the pH value to 4.0 to 9.0 with acetic acid, ammonia water, hydrochloric acid or potassium hydroxide.

4. The salt exchange method according to claim 1, characterized in that In step S2), the mass of the sample is 1.0% to 10.0% of the mass of the reverse phase chromatography filler; Preferably, the mass of the sample is 3.0% to 8.0% of the mass of the reverse phase chromatography filler; Preferably, the reverse phase chromatography filler is any one of C18, C8 or C4; Preferably, the particle size of the filler in the preparation column is 5 μm to 10 μm, and the pore size of the filler is ~30nm.

5. The salt exchange method according to claim 1, characterized in that The concentration of acetic acid in the mobile phase A2 is 0.02% to 1%.

6. The salt exchange method according to claim 1, characterized in that The mobile phase ratio used for equilibrating the preparative column in step S1) is 90%-95% mobile phase A1 and 10%-5% mobile phase B by volume.

7. The salt exchange method according to claim 1, characterized in that The mobile phase ratio used for flushing the preparation column in step S3) is 90%-95% mobile phase A1 and 10%-5% mobile phase B by volume.

8. The salt exchange method according to claim 1, characterized in that The mobile phase ratio used for flushing the preparation column in step S4) is 90%-95% mobile phase A2 and 10%-5% mobile phase B by volume.

9. The salt exchange method according to claim 1, characterized in that The mobile phase ratio used for eluting the preparative column in step S5) is 30% to 60% of mobile phase A2 and 70% to 40% of mobile phase B by volume.

10. The salt-exchanged telpotide prepared by the salt-exchanging method according to any one of claims 1 to 9; Preferably, the solubility of the salt-exchanged telpotide is above 100 mg / mL, and the pH after dissolution is 5-8.

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