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One-step method based solid-phase polypeptide synthesis method

A solid-phase synthesis and one-step technology, which is applied in the field of one-step solid-phase synthesis of peptides, can solve problems such as inconvenient operation, complicated preactivation process, and easy racemization

Active Publication Date: 2013-10-30
SPH NO 1 BIOCHEM & PHARMA CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Therefore, the technical problem to be solved by the present invention is to overcome the need to pre-activate the amino acid at low temperature in order to suppress the racemization of the protected amino acid in the condensation reaction in the existing solid-phase synthesis of the polypeptide, and then transfer the activated protected amino acid to the condensation reactor B for condensation Reaction, the preactivation process is complex, requires multi-step operation, inconvenient operation, and easy to cause racemization

Method used

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  • One-step method based solid-phase polypeptide synthesis method
  • One-step method based solid-phase polypeptide synthesis method
  • One-step method based solid-phase polypeptide synthesis method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0113] Example 1: Preparation of Fmoc-Phe-Wang resin

[0114] Weigh 50 g of Wang resin (100-200 mesh, 1.23 mmol / g), put it into a sand core reactor, wash once with 500 mL of DMF, drain, and then use 500 mL of DCM to fully swell the resin and drain. Add Fmoc-Phe-OH (MW: 387.4, 3 times the moles of Wang resin) 71.5g, HOBt (MW: 135.13, 3 times the moles of Wang resin) 24.9g, DIC (MW: 126.2, the moles of Wang resin) 3 times) 28.9mL, 300mL DMF, 100mL DCM, fully and uniformly contact the peptide resin with nitrogen flow, and use the volatilization endotherm of DCM to rapidly cool the entire reaction system to 16 ° C, slowly add DMAP (MW: 122, Wang resin moles) 0.2 times the number) 1.5 g, and the mixture was stirred at 26° C. for 18 hours. Vacuum dry, wash twice with DMF, and vacuum dry. Add Pyridine (MW: 79.1, 10 times the mole number of Wang resin) 49mL, Ac 2 O (MW: 102.09, 10 times the mole number of Wang resin) 58 mL, 500 mL DCM, the mixture was stirred at 27°C for 3 hours, d...

Embodiment 2

[0115] Example 2: Preparation of Fmoc-Pro-Phe-Wang resin

[0116] Add 500 mL of 20% Pip / DMF solution, stir at 25 °C for 10 minutes, vacuum dry, add 500 mL of 20% Pip / DMF solution, stir at 25 °C for 20 minutes, vacuum dry, wash once with DMF and twice with MeOH, DMF was washed once, DCM once, and dried in vacuo. Add Fmoc-Pro-OH (MW: 337.4, twice the moles of Fmoc-Phe-Wang resin) 40.5g, HOBt (MW: 135.13, twice the moles of Fmoc-Phe-Wang resin) 16.2g, HBTU (MW : 379.25, twice the moles of Fmoc-Phe-Wang resin) 45.5g, 350mL DMF, 100mL DCM, fully and uniformly contact the peptide resin with nitrogen flow, and use the volatilization endotherm of DCM to quickly cool the entire reaction system to 15 ℃ , 31.4 mL of DIPEA (MW: 129.24, twice the moles of Fmoc-Phe-Wang resin) was added dropwise, and the mixture was stirred at 25°C for 2.5 hours. Wash twice and vacuum dry.

Embodiment 3

[0117] Example 3: Preparation of Fmoc-His(Trt)-Pro-Phe-Wang resin

[0118] Add 500 mL of 20% Pip / DMF solution, stir at 25 °C for 10 minutes, vacuum dry, add 500 mL of 20% Pip / DMF solution, stir at 25 °C for 20 minutes, vacuum dry, wash once with DMF and twice with MeOH, DMF was washed once, DCM once, and dried in vacuo. Add Fmoc-His(Trt)-OH (MW: 619.7, twice the moles of Fmoc-Phe-Wang resin) 74.4g, HOBt (MW: 135.13, twice the moles of Fmoc-Phe-Wang resin) 16.2g, HBTU (MW: 379.25, twice the moles of Fmoc-Phe-Wang resin) 45.5g, 350mL DMF, 100mL DCM, fully and uniformly contact the peptide resin with nitrogen flow, and use the volatilization endotherm of DCM to rapidly cool down the entire reaction system To 18°C, 31.4 mL of DIPEA (MW: 129.24, twice the mole number of Fmoc-Phe-Wang resin) was added dropwise, the mixture was stirred at 23°C for 2 hours, the ninhydrin test was negative, vacuum dried, and washed once with MeOH , washed twice with DMF, and vacuum-dried.

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Abstract

The invention discloses a one-step method based solid-phase polypeptide synthesis method. The technology of blowing in nitrogen or insert gases to volatilize a volatile solvent and reduce the temperature is adopted in each condensation reaction of connected protected amino acids, the nitrogen or the insert gases are filled into a reaction mixture so that the volatile solvent is volatilized and the temperature of a reaction system can be maintained at 15-20 DEG C, after an auxiliary condensing agent is added to start the condensation reaction, the temperature of the condensation reaction system is always stabilized at 22-28 DEG C by a heat insulation device, and during condensation of protected amino acids, the amino acids are unnecessary to be activated at low temperature in an activator and then be transferred to a condensation reactor, thus all the chemical reactions for synthesizing polypeptide are continuously completed in the same reactor in sequence. The method for synthesizing angiotensinamide is environment-friendly and efficient, has low requirements for equipment and can be applied to large-scale industrial production.

Description

technical field [0001] The invention belongs to the field of pharmaceutical synthesis technology. More specifically, the present invention relates to a method for one-step solid-phase synthesis of polypeptides. Background technique [0002] There are two main methods for chemical synthesis of peptides: liquid-phase synthesis and solid-phase synthesis. The advantage of liquid-phase synthesis is that the synthesis of intermediate products in each step can be directly controlled and purified. The solid-phase synthesis process is simple, occupies less man-hours and manpower, and saves equipment and space due to less material transfer; the disadvantage is that the intermediate products in each step cannot be purified, and the reaction conditions of each step must be optimized to make the conversion rate close to 100%, and as far as possible Avoid side effects. Patent US2978444 describes a method for liquid phase synthesis of vasopressin. This process route uses a polymeric syn...

Claims

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Application Information

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IPC IPC(8): C07K1/06C07K1/04
Inventor 黄臻辉丁金国江锡铭洪勇霍建丽琚姝
Owner SPH NO 1 BIOCHEM & PHARMA CO LTD
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