Solid phase synthesis method for adrenomedullin (27-52)

An adrenomedullin and solid-phase synthesis technology, which is applied in the field of solid-phase synthesis of adrenomedullin, can solve the problems of shortening the service life of the equipment, generating more side reaction products, unstable side chain protecting groups, etc., and achieving easy purification. , The production of side reaction products is less, and it is easy to monitor and control the effect of the reaction

Inactive Publication Date: 2008-04-23
WEST CHINA HOSPITAL SICHUAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] 1. During the synthesis process, it is necessary to repeatedly use acid to remove the protecting group, and then use hydrogen fluoride to crack the peptide from the resin, so the reaction conditions are severe, which will inevitably cause side reactions, and make some of the side chain protecting groups ineffective. Stable, resulting in the gradual loss of the extended peptide chain from the solid support, low yield, and changes in the side chains of amino acid residues such as Asp (aspartic acid), Lys (lysine), etc.
[0008] 2. Due to the production of more side reaction products, the difficulty of purification is increased
[0009] 3. Since the hydrogen fluoride used is a strong acid and highly corrosive, it not only shortens the service life of the equipment, but also pollutes the environment

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] This embodiment is the first peptide grafted resin.

[0027]Weigh 0.25mmol (234mg) p-alkoxybenzyl alcohol resin and place it in a solid-phase reactor, soak it with dichloromethane for 2h and filter it with suction, then add 1mmol Fmoc-Ala-OH, 1mmol dicyclohexylcarbodiimide , 0.1mmol 4-dimethylaminopiperidine was dissolved in 6mL of N-methylpyrrolidone, added to a solid-phase reactor and reacted with p-alkoxybenzyl alcohol resin at room temperature for 2h, after the reaction was completed, dichloromethane Wash 5 times with 3ml each time, then add 4ml 20% piperidine / dimethylformamide mixed solution, shake for 0.5h, filter with suction, wash 5 times with 2ml each time of N-methylpyrrolidone, dichloromethane each time Wash 5 times with 2ml each time and wash 5 times with 3ml methanol each time, and repeat the above washing-deprotection-washing twice, and finally the reactant is drained to constant weight.

Embodiment 2

[0029] This example is the synthesis of the second peptide.

[0030] First, add 1-hydroxybenzotriazole, 1-hydroxy-benzo-triazole tetrazole prepared in 1:1:1 according to the molar ratio of the resin to the solid-phase reactor with the resin obtained in the previous step reaction. Methyl hexafluorophosphate, diisopropylethylamine condensation reagent 2mL (concentration 0.45mol / L) and 0.5mmol Fmoc-His(Trt)-OH, and add 6ml N-methylpyrrolidone, condense at room temperature for 2h, Then wash with 3ml of dichloromethane for 5 times, then add 4ml of 20% piperidine / dimethylformamide mixed solution, shake for 0.5h, filter with suction, and wash with 2ml of N-methylpyrrolidone for 5 times times, dichloromethane each 2ml wash 5 times and methanol each 3ml wash 5 times, and repeat the above washing-deprotection-washing twice, and finally the reactant was drained to constant weight.

Embodiment 3～26

[0032] This part of the examples is based on Example 2 to continue the synthesis of the third peptide to the twenty-sixth peptide.

[0033] Since the synthetic procedures and control conditions of these examples are completely the same as those of Example 2 except for the addition of protected amino acids, the description is omitted here. It is worth noting that the added protected amino acids are all 0.5 mmol, and the added protected amino acids are as follows:

[0034] Fmoc-Gln(Trt)-OH;

[0035] Fmoc-Ile-OH;

[0036] Fmoc-Tyr(tBu)-OH;

[0037] Fmoc-Gln(Trt)-OH;

[0038] Fmoc-Phe-OH;

[0039] Fmoc-Thr(tBu)-OH;

[0040] Fmoc-Asp(OtBu)-OH;

[0041] Fmoc-Lys-(Boc)-OH;

[0042] Fmoc-Asp(OtBu)-OH;

[0043] Fmoc-Lys-(Boc)-OH;

[0044] Fmoc-Asp(OtBu)-OH;

[0045] Fmoc-Asn(Trt)-OH;

[0046] Fmoc-Val-OH;

[0047] Fmoc-Ala-OH;

[0048] Fmoc-Pro-OH;

[0049] Fmoc-Arg(Pbf)-OH;

[0050] Fmoc-Ser(tBu)-OH;

[0051] Fmoc-Lys-(Boc)-OH;

[0052] Fmoc-Ile-OH;

[0053] Fmoc-Ser...

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PUM

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Abstract

The present invention discloses solid phase synthesis process of adrenal medullarin (27-52). The synthesis process includes the first solid phase Fmoc polypeptide synthesis to graft protective amino acid Fmoc-Ala-OH onto resin, connecting the rest protective amino acids successively, cutting down the synthesized coarse peptide with peptide cutting reagent, and final separating and purifying to obtain the adrenal medullarin (27-52) as the target peptide. The present invention has mild reaction condition, high reaction rate, less side reactions, easy purification, high yield, no corrosion to the equipment and no environmental pollution.

Description

technical field [0001] The invention belongs to the technical field of solid-phase synthesis method and application of polypeptide, and in particular relates to a solid-phase synthesis method of adrenomedullin (27-52). Background technique [0002] Adrenomedullin (ADM) is expressed in the normal human adrenal gland, and it is also highly expressed in bone tissue and secreted by osteoblasts. [0003] Adrenomedullin is a potent stimulator of osteoblastic activity invitro and in vivo. Am J Physiol 1997; 273: E1113-20 ) found that its homology with the calcitonin gene-related peptide (CGRP), a drug for treating osteoporosis, is 24%, and should belong to the CGRP family; human ADM is composed of Composed of 52 amino acid residues, the 16th and 21st cysteine ​​residues constitute a ring structure of disulfide bonds in the chain. This structural feature is very important for its biological activity, but it suggests that the ring structure is not responsible for Necessary for oste...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/575C07K1/04
CPCY02P20/55
Inventor 张晖陈治宇裴福兴王怡鑫
Owner WEST CHINA HOSPITAL SICHUAN UNIV
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