A method for splitting thymosin β4

A technology of thymosin and lysate, which is applied in the field of peptide cleavage, can solve the problems of large amount of reagents used, environmental pollution, unsuitable for thymosin β4, etc., and achieves the effect of facilitating industrial production, reducing production costs, and improving the purity of crude peptides

Active Publication Date: 2015-10-21
哈尔滨吉象隆生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0010] Method 1 requires high equipment requirements, high cost investment, high risk of operation, and serious environmental pollution
[0011] Method 2 has a cumbersome operation process, and the operation is dangerous, and the amount of reagent used is large, which increases the cost of the product
[0012] Due to the particularity of thymosin β4, the above method is not suitable for the cleavage of thymosin β4. After research, the present invention has found a new cleavage method, which is especially suitable for the cleavage of thymosin β4.

Method used

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  • A method for splitting thymosin β4
  • A method for splitting thymosin β4
  • A method for splitting thymosin β4

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] Thymosin β4 peptide resin was obtained by solid phase synthesis.

[0064] Prepare a lysate with a mass ratio (trifluoroacetic acid: thioanisole: ethanedithiol: phenol: water = 86:4:3:4:3) of 100 grams, and stir the prepared lysate evenly. Weigh 5% thymosin β4 peptide resin (5 g), add to the above lysate, and react for 120 minutes. After the reaction is completed, filter the above reaction solution with a sand core funnel, collect the filtrate, wash the filter cake with trifluoroacetic acid, and combine the filtrate. The filtrate was added to 1000 ml of glacial ether for precipitation, filtered with a sand core funnel, the filter cake was collected, washed with glacial ether, and put into a vacuum drying oven for vacuum drying to obtain thymosin β4 crude peptide. Weighed to get 2.55 grams of crude product, crude peptide weight yield: 96.2%, through HPLC detection, crude peptide purity: 29.3%

[0065] Purified by high performance liquid phase to obtain thymosin β4 fine ...

Embodiment 2

[0067] Thymosin β4 peptide resin was obtained by solid phase synthesis.

[0068] Prepare lysate, mass ratio (trifluoroacetic acid: thioanisole: ethanedithiol: phenol: water: methyl sulfide = 84: 4: 3: 4: 3: 2) 100 grams, stir the prepared lysate uniform. Weigh 5% thymosin β4 peptide resin (5 g), add to the above lysate, and react for 120 minutes. After the reaction is completed, filter the above reaction solution with a sand core funnel, collect the filtrate, wash the filter cake with trifluoroacetic acid, and combine the filtrate. The filtrate was added to 1000 ml of glacial ether for precipitation, filtered with a sand core funnel, the filter cake was collected, washed with glacial ether, and put into a vacuum drying oven for vacuum drying to obtain thymosin β4 crude peptide. Weighed 2.63 grams of crude product, crude peptide weight yield: 99.2%, through HPLC detection, crude peptide purity: 33.5%

[0069] Purified by high performance liquid phase to obtain thymosin β4 fi...

Embodiment 3

[0071] Thymosin β4 peptide resin was obtained by solid phase synthesis.

[0072] Configure lysate, mass ratio (trifluoroacetic acid: thioanisole: ethanedithiol: phenol: water: methyl sulfide: ammonium iodide = 83.5: 4: 3: 4: 3: 2: 0.5) 100 grams, will The prepared lysate was stirred evenly. Weigh 5% thymosin β4 peptide resin (5 g), add to the above lysate, and react for 120 minutes. After the reaction is completed, filter the above reaction solution with a sand core funnel, collect the filtrate, wash the filter cake with trifluoroacetic acid, and combine the filtrate. The filtrate was added to 1000 ml of glacial ether for precipitation, filtered with a sand core funnel, the filter cake was collected, washed with glacial ether, and put into a vacuum drying oven for vacuum drying to obtain thymosin β4 crude peptide. Weighed 2.88 grams of crude product, crude peptide weight yield: 108.7%, tested by HPLC, crude peptide purity: 43.5%

[0073] Purified by high performance liquid ...

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Abstract

The invention discloses a method for pyrolysis of thymosin beta-4. The method comprises the step of carrying out pyrolysis on thymosin beta-4 peptide resin to obtain crude peptide, wherein a mixed solvent of trifluoroacetic acid, thioanisole, dithioglycol, phenol, water, dimethyl sulfide and ammonium iodide is used as a pyrolysis reagent in the pyrolysis process. According to the method for pyrolysis of thymosin beta-4, the purity of crude peptide is greatly improved, meanwhile the total yield is greatly increased, the production cost is reduced, and the method is simple in process and convenient for industrial production.

Description

technical field [0001] The invention relates to a method for cleavage of polypeptides, in particular to a method for cleavage of thymosin β4. Background technique [0002] Thymosin was first extracted from calf thymus by Goldstein and White in 1966. According to the different isoelectric points, thymosin can be divided into 3 types: α (PI<5), β (5<PI<7) and γ (PI>7). Constructs have been identified, and there are three main types in the human body, namely thymosin β4 (Tβ4), Tβ10 and Tβ15, of which thymosin β4 has the highest content. Thymosin β4 is a highly conserved water-soluble polypeptide composed of 43 amino acid residues, with a relative molecular mass of 4964. [0003] The molecular formula of thymosin β4 is: [0004] Ac-Ser-Asp-Lys-Pro-Asp-Met-Ala-Glu-Ile-Glu-Lys-Phe-Asp-Lys-Ser-Lys-Leu-Lys-Lys-Thr-Glu-Thr-Gln-Glu -Lys-Asn-Pro-Leu-Pro-Ser-Lys-Glu-Thr-Ile-Glu-Gln-Glu-Lys-Gln-Ala-Gly-Glu-Ser-OH Thymosin β4 is considered to be predominantly spherical Ac...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/575C07K1/04
CPCC07K14/57581
Inventor 冷国庆余荣熹王艳王浩迟帅刘宇帅田辉张琪李迎新肖明辉李喜全赵民喜苏宏健
Owner 哈尔滨吉象隆生物技术有限公司
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