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Gonadorelin purification method

A volume percentage and gradient technology, applied in the field of gonadorelin purification, can solve the problems of low purity of crude peptide, low yield, difficult product control, etc., and achieve the effect of reducing cost, increasing sample loading, and convenient operation

Inactive Publication Date: 2016-06-08
GL BIOCHEM SHANGHAI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to provide a method for purifying gonadorelin suitable for large-scale industrial production, which mainly solves the problems of low purity of crude peptide in the production process of the product, difficult control of the product, and low yield.

Method used

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Effect test

Embodiment 1

[0018] 1. Sample Processing

[0019] Weigh 5 g of crude gonadorelin and add 5% acetonitrile by volume to dissolve it ultrasonically until the liquid is clear, filter through a filter membrane with a pore size of 0.45 um, and collect the filtrate for later use.

[0020] 2. Primary purification

[0021] Chromatographic column: SPE column. Purification process: put the above filtrate on the SPE column, the SPE column is HyperSepPEP, first eluted with 10% acetonitrile + 0.1% TFA aqueous solution by volume, and then eluted with 40% acetonitrile + 0.1% TFA aqueous solution by volume, Collect the peptide solution of the peak of interest. Concentrate the collected fractions to 40-50 mg / mL by rotary evaporation under reduced pressure.

[0022] 3. Secondary purification

[0023] Chromatographic column: C18 chromatographic column, column diameter and length: 50cm×25cm. Mobile phase: mobile phase A is 0.01-0.5% by volume phosphoric acid aqueous solution, mobile phase B is chromatograp...

Embodiment 2

[0027] 1. Sample Processing

[0028] Weigh 15g of crude gonadorelin and add 5% acetonitrile by volume to dissolve it ultrasonically until the liquid is clear, filter through a filter membrane with a pore size of 0.45um, and collect the filtrate for later use.

[0029] 2. Primary purification

[0030] Chromatographic column: SPE column. Purification process: put the above filtrate on the SPE column, which is an MCXSPE column, first elute with 10% acetonitrile + 0.1% TFA aqueous solution by volume, then elute with 40% acetonitrile + 0.1% TFA aqueous solution by volume, and collect Peptide solution of the target peak. Concentrate the collected fractions to 40-50 mg / mL by rotary evaporation under reduced pressure.

[0031] 3. Secondary purification

[0032] Chromatographic column: C18 chromatographic column, column diameter and length: 100cm×25cm. Mobile phase: mobile phase A is 0.01-0.5% by volume phosphoric acid aqueous solution, mobile phase B is chromatographically pure ac...

Embodiment 3

[0036] 1. Sample Processing

[0037] Weigh 25g of crude gonadorelin and add 5% acetonitrile by volume to dissolve it ultrasonically until the liquid is clear, filter through a filter membrane with a pore size of 0.45um, and collect the filtrate for later use.

[0038] 2. Primary purification

[0039] Chromatographic column: SPE column. Purification process: put the above filtrate on the SPE column, the SPE column is HyperSepPEP, first eluted with 10% acetonitrile + 0.1% TFA aqueous solution by volume, and then eluted with 40% acetonitrile + 0.1% TFA aqueous solution by volume, Collect the peptide solution of the peak of interest. Concentrate the collected fractions to 40-50 mg / mL by rotary evaporation under reduced pressure.

[0040] 3. Secondary purification

[0041]Chromatographic column: C18 chromatographic column, column diameter and length: 150cm×25cm. Mobile phase: mobile phase A is 0.01-0.5% by volume phosphoric acid aqueous solution, mobile phase B is chromatograph...

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Abstract

The invention aims to provide a gonadorelin purification method applicable to large-scale industrial production and mainly solves the problems of low crude peptides, difficulty in product control, low yield and the like in a production process of products. The technical scheme is that gonadorelin is subjected to gradient elution purification through an SPE column and a reversion phase chromatographic column, gonadorelin fractions with higher purity are obtained, the gonadorelin fractions are converted into salt with an HPLC conversion-into-salt method, reduced-pressure rotary evaporation, concentration, freezing and drying are performed, and gonadorelin acetate is obtained. The method is used for industrially purifying gonadorelin.

Description

technical field [0001] The invention relates to a method for purifying gonadorelin, in particular to a method for purifying gonadorelin with low cost, less pollution and high efficiency, which is suitable for industrial purification. Background technique [0002] Gonadorelin, whose chemical name is 5'-oxoprolyl-L-histidyl-L-tryptophanyl-L-seryl-L-tyrosyl-glycyl-L-arginine Acyl-L-arginyl-L-prolyl-glycylamide, its structural formula: [0003] [0004] Gonadorelin can stimulate the synthesis and release of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in the anterior pituitary gland, thereby detecting the reserve function of pituitary gonadotropins. After injection of gonadorelin in normal people, the increase of LH is obvious, which is higher than that of FSH, and the FSH response of prepubertal women is higher than that of LH; those with insufficient gonadotropin-releasing hormone may have a delayed reaction after injection, and sometimes need intravenou...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/23C07K1/20
CPCC07K7/23
Inventor 徐红岩秦敬国
Owner GL BIOCHEM SHANGHAI
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