Solid-phase synthesis method of bremelanotide

A technology of bremelanotide and solid-phase synthesis, which is applied to the preparation method of peptides, chemical instruments and methods, and peptides, and can solve problems such as difficult ring formation and large steric hindrance

Inactive Publication Date: 2016-05-25
JINAN KANGHE MEDICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Ring formation in the liquid phase is easy to produce a large number of intermolecular coupling products, and the solid phase ring formation method is difficult due to too much steric hindrance

Method used

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  • Solid-phase synthesis method of bremelanotide
  • Solid-phase synthesis method of bremelanotide
  • Solid-phase synthesis method of bremelanotide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] Example 1: HO-(CH 2 ) 7 - Preparation of CONH-RinkAmideResin

[0071] Accurately weigh 100g of RinkAmideResin with a substitution degree of 0.52mmol / g and place it in a solid-phase synthesis kettle, wash twice with DMF, 500ml / time, 3min / time, after washing, add 500mlDCM to swell the resin for 30min.

[0072] RinkAmideResin was deprotected with 20% piperidine / DMF, 400ml / time, the first reaction was 5min, and the second reaction was 15min. After deprotection twice, the resin was washed six times with DMF, 500ml / time, 3min / time. Take a small sample of the resin and detect it with ninhydrin reagent, the resin is dark blue.

[0073] Weigh HO-(CH 2 ) 7 -COOH24.9g (156mmol, 3eq), HOBt23.2g (171.6mmol, 3.3eq) were placed in a dissolving bottle, dissolved in 200ml of DMF, pre-cooled at 0-5°C for 10min, and activated by adding DIC27.5ml (171.6mmol, 3.3eq) 5min, put into the resin after the activation is completed, stir and react at 25°C for 2h. After 2 hours, the resin was...

Embodiment 2

[0074] Example 2: HO-(CH 2 ) 15 - Preparation of CONH-RinkAmideResin

[0075] Accurately weigh 100g of RinkAmideResin with a substitution degree of 0.52mmol / g and place it in a solid-phase synthesis kettle, wash twice with DMF, 500ml / time, 3min / time, after washing, add 500mlDCM to swell the resin for 30min.

[0076] RinkAmideResin was deprotected with 20% piperidine / DMF, 400ml / time, the first reaction was 5min, and the second reaction was 15min. After deprotection twice, the resin was washed six times with DMF, 500ml / time, 3min / time. Take a small sample of the resin and detect it with ninhydrin reagent, the resin is dark blue.

[0077] Weigh HO-(CH 2 ) 15 -COOH42.4g (156mmol, 3eq), HOBt23.2g (171.6mmol, 3.3eq) was placed in a dissolving bottle, dissolved in 200ml of DMF, pre-cooled at 0-5°C for 10min, and activated by adding DIC27.5ml (171.6mmol, 3.3eq) 5min, put into the resin after the activation is completed, stir and react at 25°C for 2h. After 2 hours, the resin wa...

Embodiment 3

[0078]Example 3: Fmoc-Lys(Mtt)-O-(CH 2 ) 7 - Preparation of CONH-RinkAmideResin

[0079] Accurately weigh Fmoc-Lys(Mtt)-OH97.344g (156mmol, 3eq), HOBt23.2g (171.6mmol, 3.3eq) into a dissolving bottle, dissolve with 300ml DMF, pre-cool at 0-5°C for 10min, add DIC27. 5ml (171.6mmol, 3.3eq) activation 5min, drop into the synthetic resin HO-(CH of embodiment 1 after activation completes 2 ) 7 -CONH-RinkAmideResin, after stirring for 10 minutes, add DMAP 1.9g (15.6mmol, 0.3eq), and stir for 5 hours at 25°C. After 5h, the resin was washed and dried in a vacuum drying phase (<-0.1Mpa, 8h). A sample was taken to measure the degree of substitution, and the measured value was 0.33mmol / g.

[0080] Weigh 90g of the above dry resin (substitution degree is 0.33mmol / g, 30mmol) and place it in a solid-phase synthesis kettle, wash twice with DMF, 500ml / time, 3min / time, add 500mlDCM to swell the resin for 30min after washing.

[0081] Measure 45ml (450mmol, 15eq) of acetic anhydride and 3...

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Abstract

The invention belongs to the field of polypeptide synthesis, and particularly relates to a synthesis method using RinkAmide Resin as a solid-phase carrier and HO-(CH2)n-COOH as Linker for connecting a solid-phase carrier and a target peptide. In the method, the carboxyl terminal is connected with a solid-phase carrier by an amido bond, and the hydroxyl terminal is connected with the target peptide by virtue of an ester bond. Compared with the prior art, according to the technical scheme, long-chain fatty acid is introduced between the solid-phase carrier and bremelanotide, the space environment of target peptide solid-phase cyclization is greatly optimized, intermolecular coupling occurring during liquid-mode cyclization can be avoided, and the purity of the final target peptide-Linker is over 85 percent, the total yield of purification is over 50 percent, and the fine peptide purity is over 99 percent.

Description

technical field [0001] The invention relates to the field of polypeptide synthesis, in particular to a method for solid-phase synthesis of bremelanotide by using long-chain fatty acids as Linker. Background technique [0002] Bremelanotide, the English name is Bremelanotide, also known as Bremelanotide, PT141, which consists of 7 amino acids and has the same core sequence as α-melanocyte-stimulating hormone, whose receptor is in appetite and sexual desire arousal makes an important impact. Its peptide sequence is AC-Nle-[Asp-His-D-Phe-Arg-Trp-Lys] cycle -OH, the structural formula is as follows: [0003] [0004] Sexual dysfunction is divided into male sexual dysfunction and female sexual dysfunction. Most of the drugs currently used to treat male sexual dysfunction are phosphodiesterase 5 (PDE-5) inhibitors, such as cetenafil and tadalafil , vardenafil. The mechanism of action of these drugs is to maintain or increase the concentration of cyclic adenosine monophospha...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/56C07K1/10C07K1/06C07K1/04
CPCY02P20/55C07K7/56
Inventor 张颖陈雷王仁友李同金石鑫磊
Owner JINAN KANGHE MEDICAL TECH
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