Method for synthesizing cholecystokinin octapeptide by combining solid phase method and liquid phase method

A technology of cholecystokinin and solid-phase method, which is applied in the preparation of trifluoroacetate), cholecystokinin octapeptide (sincalide including acetate), which can solve the troublesome separation of intermediate products and easily produce by-products , long preparation cycle and other issues, to achieve the effect of easy post-processing, considerable economic and practical value, and low cost

Active Publication Date: 2012-11-14
GL BIOCHEM SHANGHAI +2
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0012] The purpose of the present invention is to provide a method for the synthesis of octapeptide cholecystokinin by combining a solid-phase method and a liquid-phase method, which mainly solves the troublesome separation of intermediate products in the existing synthesis method, the long preparation cycle and the easy production of by-products in the reaction, and the high cost , low yield and other technical problems

Method used

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  • Method for synthesizing cholecystokinin octapeptide by combining solid phase method and liquid phase method

Examples

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Effect test

Embodiment 1

[0038] Example 1 Synthesis by liquid phase method: H-Asp(otbu)-Phe-NH 2

[0039] Take 265g of Boc-Phe-OH and pour it into a round bottom flask, then add 300ml of THF (tetrahydrofuran) to dissolve it, add 133ml of NMM, at low temperature, slowly add 114ml of ClCOOET dropwise, and keep stirring. Filter out the precipitate, then collect the filtrate, add 77ml of ammonia water dropwise to the filtrate, react at room temperature for 40min, add 250ml of water, evaporate THF to dryness, then add 250ml of ethyl acetate to extract the target product, extract 3 times, then spin the filtrate to dryness, Add 300ml TFA to remove Boc, room temperature for 1 hour, add 1500ml glacial ether to precipitate, filter, dry in a positive air dryer to obtain a solid: H-Phe-NH 2 , 152g, H-Phe-NH 2 Pour into a round bottom flask, add 379g Fmoc-Asp(otbu)-OH, then dissolve with 200ml DMF, add 17.3ml DIC, 15g HOBT. Stir and react overnight at room temperature, add 250ml ethyl acetate to extract after c...

Embodiment 2

[0040] Example 2 Preparation of Fmoc-Met-CTC Resin

[0041] Dissolve 25g of Fmoc-Met-OH with 200ml of DCM and add it to a 2000ml solid-phase reactor, then add 50g of CTC Resin, then add 60ml of DIEA dropwise, blow the reaction with nitrogen for 2 hours, and then add 500ml of methanol Block for 30 minutes, filter the amino acid resin with a Buchner funnel, then wash and shrink three times with DMF, DCM, and methanol;

[0042] CTC Resin with a substitution degree of 0.3mmol / g was used as the original resin for reaction. The obtained Fmoc-Met-CTC Resin detects that the substitution degree is 0.2mmol / g. In this way, the production process with the same number of moles consumes resin, and the equivalent amount of solvent is large, which is uneconomical and not conducive to environmental protection. It is not suitable for use; CTC Resin with a substitution degree of 1.5mmol is used Carry out the reaction to obtain Fmoc-Met-CTC Resin. The detection degree of substitution is 0....

Embodiment 3

[0043] Example 3 Preparation of Fmoc-Met-CTC Resin

[0044] Choose CTC Resin with a substitution degree of 1.0mmol / g, the reaction needs to add organic base N,N-diisopropylethylamine, and the rest are the same as in Example 2, and finally the substitution degree of Fmoc-Met-CTC Resin is: 0.75 mmol / g.

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Abstract

The invention relates to a preparation method of cholecystokinin octapeptide, in particular to a method for synthesizing the cholecystokinin octapeptide by combining a solid phase method and a liquid phase method. The method mainly solves the technical problem that the existing synthesis method is troublesome in separation of intermediate products, long in preparation period, apt to produce by-products in reaction, high in cost, low in yield and the like. The technical scheme is that the synthesis method comprises the following steps of: (1) synthesizing L-aspartyl-4-tertiary butyl ester-benzene propanamide by using the liquid phase method; (2) synthesizing cholecystokinin octapeptide full-protection fragments by using the solid phase method; (3) carrying out weak acid cutting on the full-protection fragments; (4) carrying out liquid phase condensation on the full-protection fragments and dipeptide fragments to obtain full-protection cholecystokinin octapeptide; (5) cutting, adding the full-protection cholecystokinin octapeptide in cutting fluid for cutting, and then adding ice diethyl ether for sediment to obtain cholecystokinin octapeptide crude products; and (6) purifying the crude products through high-phase liquid chromatogram, preparing, rotatably steaming, carrying out freeze-drying to obtain the cholecystokinin octapeptide competitive products. The method is used for preparing the cholecystokinin octapeptide.

Description

technical field [0001] The invention relates to a preparation method of octapeptide cholecystokinin (sincalide includes acetate and trifluoroacetate), in particular to a method for synthesizing octapeptide cholecystokinin by combining a solid-phase method and a liquid-phase method. Background technique [0002] Octapeptide cholecystokinin, also known as: Sincalide, English name: Sincalide [0003] Structural formula: H-ASP-TYR(SO3H)-MET-GLY-TRP-MET-ASP-PHE-NH2 [0004] [0005] Molecular formula and molecular weight: C49H62N10O16S3; 1143.27 [0006] Properties: also known as cholecystokinin, incretin, cholecystokinin, pancreatin secretin. A single-chain polypeptide hormone extracted from pig upper duodenal mucosa. Consists of 33 amino acids. A digestive hormone that stimulates the secretion and growth of the exocrine pancreas, and also plays a role in satiety, pain and nerve conduction. It is secreted by the M[I] cells of the duodenum, jejunum, and ileum. [0007] S...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/06C07K1/06C07K1/04C07K1/02
Inventor 徐红岩沈永刚
Owner GL BIOCHEM SHANGHAI
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