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Urinary follicle stimulating hormone with high purity and preparation method thereof

A follicle-stimulating hormone and follicle-stimulating hormone technology, applied in the field of protein purification and biomedicine, can solve the problems of FSH specific activity and purity not very ideal, side effects, etc.

Active Publication Date: 2009-10-14
SHANGHAI TECHWELL BIOPHARMACEUTICALS CO LTD
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  • Summary
  • Abstract
  • Description
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  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The current preparation method of urine-derived FSH is mainly based on low-purity urinary follicle-stimulating hormone or menopausal gonadotropin (HMG) as the starting material, through hydrophobic chromatography, or monoclonal antibody immunochromatography and reversed-phase HPLC. Purification, but the specific activity and purity of the obtained FSH are not very satisfactory, and there are occasional reports of side reactions

Method used

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  • Urinary follicle stimulating hormone with high purity and preparation method thereof

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preparation example Construction

[0053] The preparation method of the high-purity urogenous FSH provided by the invention comprises steps:

[0054] The low-purity urinary follicle-stimulating hormone or menopausal gonadotropin (HMG) is purified by chromatography to obtain high-purity urinary FSH; the chromatography is cation exchange chromatography and dye affinity chromatography.

[0055] Preferably, the method comprises the steps of:

[0056] (1) purifying solution 1 containing low-purity FSH through cation exchange chromatography to obtain distillate 1; and

[0057] (2) The solution 2 containing distillate 1 was purified by dye affinity chromatography to obtain high-purity urinary follicle-stimulating hormone (pFSH).

[0058] The skeleton medium of the chromatographic column of the cation-exchange chromatography used in the preparation method of the present invention comprises cross-linked products of agarose, dextran, cellulose, styrene, acrylic acid and / or derivatives; the chromatographic column of the ...

Embodiment 1

[0100] Preparation of high-purity urinary follicle-stimulating hormone I

[0101] Low-purity urinary follicle stimulating hormone (purchased from Shanghai Tianwei Bio-Pharmaceutical Co., Ltd.) was used as the starting material, wherein the biological potency of FSH was 315 IU / mg, and the biological potency of LH was ≤ 3 IU / mg.

[0102] Dissolve 10g of the above-mentioned low-purity UFFS with 300mL equilibration solution (0.03M sodium dihydrogen phosphate, pH5), and then apply it to a 250mL CM-Sepharose chromatography column (provided by Amersham), which has been previously used with the same equilibration solution. Well balanced. After loading, wash 10 times the column volume with washing solution (0.1M sodium acetate, pH5), and then perform 0-100% linear gradient elution with washing solution and eluent (0.1M sodium acetate + 1M NaCl, pH5), Monitor at 280nm with an ultraviolet detector, distribute and collect each distillate peak, detect its FSH immune titer, combine about 0...

Embodiment 2

[0108] Preparation of high-purity urinary follicle-stimulating hormone II

[0109] Dissolve 5g of low-purity urinary follicle stimulating hormone (the same starting material as in Example 1) with 150mL of equilibrium solution (0.03M sodium dihydrogen phosphate, pH4.8), and then apply it to a 130mL SP-Sepharose chromatographic column (provided by Amersham) , the column was previously equilibrated with the same equilibration solution. After loading, wash 10 times the column volume with washing solution (0.1M sodium acetate, pH4.8), and then perform 0-100% linear gradient washing with washing solution and eluent (0.1M sodium acetate + 1M NaCl, pH5). Remove (volume ratio concentration of eluent, within 2 hours), monitor 280nm place with UV detector, collect each distillate peak in distribution, detect its FSH immune titer, combine about 0.2L of active ingredients, add pre-cooled Water ethanol was precipitated overnight, and the precipitate was collected by centrifugation the next...

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Abstract

The invention discloses a urinary follicle stimulating hormone with a purity of no more than 95w / w%; and the impurity content is not more than 5w / w%. Simultaneously, the invention discloses a preparation method for the urinary follicle stimulating hormone with high purity.

Description

technical field [0001] The present invention relates to the fields of protein purification and biomedicine. Specifically, the present invention relates to high-purity follicle-stimulating hormone (FSH), a preparation method thereof, and a pharmaceutical composition containing the same. Background technique [0002] Follicle-stimulating hormone (FSH) is a hormone produced by the pituitary gland, which consists of two subunits, an alpha chain and a beta chain. The α subunit of FSH is exactly the same as the α subunit of leuteinizing hormone (LH) and chorionic gonadotropin (CG), with 92 amino acids, molecular weight of about 14500D, positions 52 and 78 The asparagine above is an amino acid that undergoes N-glycosylation. [0003] The β subunit of FSH consists of 111 amino acids with a molecular weight of about 18000D, of which the asparagine at the 7th and 24th positions are N-glycosylated amino acids. The β subunit of LH consists of 121 amino acids with a molecular weight o...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/59C07K1/22C07K1/18A61K38/24A61P15/08
CPCC07K14/59A61P15/08
Inventor 季晓铭高霄梁季斌郭照晔洪云海
Owner SHANGHAI TECHWELL BIOPHARMACEUTICALS CO LTD
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