Method for preparing ziconotide

A technology of ziconotide and solid-phase synthesis, which is applied in the field of peptide synthesis, can solve the problems of unfavorable high purity, increased impurities, unfavorable and other problems, achieve wide application prospects, improve reaction efficiency, and avoid the effect of impact

Inactive Publication Date: 2012-04-18
HYBIO PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the adoption of the multi-pair disulfide bond directional oxidation method and the use of various side chain removal and oxidation reagents, each step will inevitably bring about an increase in impurities, which is not conducive to obtaining high-purity, high-yield products. It is not conducive to the enlargement of the process

Method used

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  • Method for preparing ziconotide
  • Method for preparing ziconotide
  • Method for preparing ziconotide

Examples

Experimental program
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Effect test

Embodiment 1

[0038] Embodiment 1: the synthesis of Fmoc-Cys(Acm)-Rink Amide resin

[0039] Rink Amide resin 14.3g, substitution degree is 0.7mmol / g, joins in the solid-phase reaction column, after adding DCM to swell the resin for 30 minutes, remove Fmoc with 20% hexahydropyridine / DMF, wash 6 times with DMF, the 1.87 gFmoc-Cys(Acm)-OH, 1.71gHATU, 0.73gHOAt were dissolved in DMF under ice-cooling conditions, added to the above resin and reacted for 10min, then added 1.2ml TMP, and reacted at room temperature for 45min. After washing 3 times with DMF, 3 times with DCM, and 3 times with methanol for 3 min, 5 min and 8 min respectively, the resin was shrunk to obtain Fmoc-Cys(Acm)-RinkAmide resin, and the detected substitution degree was 0.4 mmol / g. The route is as follows:

[0040]

Embodiment 2

[0041] Embodiment 2: the synthesis of Fmoc-Cys(Acm)-Rink Amide MBHA resin

[0042] Add 13.8 g of Rink Amide MBHA resin, with a substitution degree of 0.6 mmol / g, into the solid-phase reaction column, add DCM to swell the resin for 30 minutes, remove Fmoc with 20% hexahydropyridine / DMF, wash with DMF 6 times, and 1.87g Fmoc-Cys(Acm)-OH, 1.71g HATU, 0.73g HOAt were dissolved in DMF under ice-cooling conditions, added to the above resin and reacted for 10min, then 1.2ml TMP was added and reacted at room temperature for 45min. After washing 3 times with DMF, 3 times with DCM, and 3 times with methanol for 3 min, 5 min and 8 min respectively, the resin was shrunk to obtain Fmoc-Cys(Acm)-Rink Amide MBHA resin, and the detected substitution degree was 0.35 mmol / g.

Embodiment 3

[0043] Embodiment 3: the synthesis of Fmoc-Cys(Acm)-Rink Amide BHA resin

[0044] With Rink Amide BHA resin 15.6g, substitution degree is 0.6mmol / g, joins in the solid-phase reaction column, after adding DCM swelling resin 30 minutes, remove Fmoc with 20% hexahydropyridine / DMF, wash 6 times with DMF, will 1.87g Fmoc-Cys(Acm)-OH, 1.71g HATU, 0.73g HOAt were dissolved in DMF under ice-cooling conditions, added to the above resin and reacted for 10min, then 1.2ml TMP was added and reacted at room temperature for 45min. After washing with DMF for 3 times, washing with DCM for 3 times, and using methanol for 3 minutes, 5 minutes and 8 minutes respectively, the resin was shrunk to obtain Fmoc-Cys(Acm)-Rink Amide BHA resin, and the detected substitution degree was 0.37mmol / g.

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Abstract

The invention discloses a method for preparing ziconotide. The technical scheme of the invention comprises the following steps: (1) obtaining an Fmoc-Cys(Acm)-amino resin from Fmoc-Cys(Acm)-OH and an amino resin; (2) obtaining a linear-ziconotide-amino resin of which a Cys side chain comprises Acm by performing the solid phase synthesis on the Fmoc-Cys(Acm)-amino resin and an amino acid adopting Fmoc group protection; (3) obtaining a linear crude peptide of which the Cys side chain comprises the Acm by performing cracking on the linear-ziconotide-amino resin of which the Cys side chain comprises the Acm, and obtaining linear ziconotide by removing the Acm, purifying and freeze-drying; (4) and obtaining the ziconotide by performing cyclization, purifying and freeze-drying on the linear ziconotide. The method for preparing ziconotide has the characteristics of simple reaction operation, easy subsequent treatment, low raw material investment, low cost, high yield and the like, and has considerable economic and practical value, and also has wide application prospect in the field of design synthesis of polypeptide drugs.

Description

technical field [0001] The invention relates to a synthesis method of polypeptide, especially a solid-phase synthesis method of ziconotide. Background technique [0002] Ziconotide (Ziconotide) is an N-type calcium channel (N-type calcium channel (NCC)) blocker, which is extracted from a toxin secreted by a fish-eating cone snail (Conus magus) in the South Pacific, containing 25 Amino Acids, Peptide Toxoid Synthesis of Three Pairs of Disulfide Bonds. [0003] Currently used painkillers mainly include the following two mechanisms: non-steroidal anti-inflammatory drugs (such as ibuprofen) and COX-2 inhibitors (such as Vioxx and Celebrex) work by blocking the pain-causing, COX enzymes in periarticular inflammation; narcotics (such as morphine, cocaine, and oxycodone) block pain-sensing opioid receptors in the brain for more severe pain. [0004] Opioid painkillers are currently the most effective painkillers, which can relieve any type of pain, but their use is greatly limite...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/435C07K1/06C07K1/04C07K1/02
Inventor 宓鹏程李红玲马亚平袁建成
Owner HYBIO PHARMA
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