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Antineoplastic cytotoxin of snake venom and preparing technique thereof

A technology of cytotoxin and production process, which is applied in the direction of antineoplastic drugs, peptide preparation methods, animal/human peptides, etc. It can solve the problems of enhanced hemolysis, interference with CTX biological activity, increased cytotoxicity, etc., and achieves good repeatability , Good process amplification ability, high resolution effect

Inactive Publication Date: 2008-09-24
FUJIAN MEDICAL UNIV
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

However, with the in-depth study of cytotoxins, many scholars have noticed that traces of phospholipase A will appear in traditional separation methods. 2 (PLA 2 ) (0.1~0.5%, w / w) co-eluted with CTX, while PLA 2 It will have a synergistic effect with CTX, which can significantly enhance the hemolysis of CTX and increase its cytotoxicity several times, which seriously interferes with the research on the biological activity and mechanism of CTX, so the isolated PLA-free 2 CTX is important

Method used

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  • Antineoplastic cytotoxin of snake venom and preparing technique thereof
  • Antineoplastic cytotoxin of snake venom and preparing technique thereof
  • Antineoplastic cytotoxin of snake venom and preparing technique thereof

Examples

Experimental program
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Embodiment 1

[0020] 1. Ion-exchange chromatography of crude cobra venom:

[0021] Dissolve 10.0 g of Zhoushan cobra venom crude venom in 100 ml of double distilled water, overnight at 4°C; centrifuge at 15,000×g at low temperature (4°C) for 15 minutes, put on SP Sepharose High Performance cation exchange chromatography column (5.0×29cm), and use 0.05 mol / L phosphate buffer (pH 5.8) and 0.0-0.5mol / L NaCl step gradient elution, 10ml / tube, flow rate 18ml / min, use The protein purification system monitors and collects elution peaks. The SP Sepharose H.P column chromatographic separation of the crude venom of cobra venom: totally obtain 20 protein peaks (see figure 1 ).

[0022] 2. Identification of biological activity of each isolated component

[0023] 1) Effect of components on rat isolated heart perfusion

[0024] Referring to the Langendorff method, the healthy SD rats were killed and the heart was quickly taken out (the heart was connected to the aorta with a length of about 1 cm), an...

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Abstract

The present invention discloses an anti-tumour snake venom cytotoxin and a production technique thereof, belonging to the fields of biochemistry and biological medicines. The snake venom cytotoxin of the present invention is derived from the prior lyophilized cobra venom powder. In the production technique of the present invention, the lyophilized cobra venom powder is resolved in the known buffer solution, and then on a SP-Sepharose H.P cation exchange column (5.0cm multipled by 29cm), gradiently eluted stage by stage by phosphate buffer and NaC1 at 10ml per tube and the flowing speed of 18ml per minute, a KTA Explorer protein purification system is used to monitor and collect eluting peaks, XII and XIII peaks with cytotoxic effects are collected, and via Source 30RPC reversed phase chromatography, cytotoxin XII and cytotoxin XIII, the molecular weight of which is about 6000 daltons to 7000 daltons and the isoelectric point of which is larger than 10, are respectively obtained. Compared with the purification step of the prior art, the production technique for producing the pure anti-tumour snake venom cytotoxin product without PLA2 has good rapid repeatability and is suitable for expanded production.

Description

technical field [0001] The invention relates to the fields of biochemistry and biomedicine, in particular to a snake venom cytotoxin and a production process thereof. Background technique [0002] The separation and purification of cytotoxin was first used by Indian scholars in 1947 by the salting-out method (Na 2 SO 4 and NaCl) from Indian cobra venom; then Ravdonat and Holler used paper chromatography; Braganca et al. separated CytotoxinP through CM-cellulose ion exchange chromatography, Sephadex G-50 gel filtration and other steps 6 ; Du Yucang et al. used SP-Sephadex C-25 cation exchange chromatography to separate 5 cytotoxins from Chinese cobra venom. However, with the in-depth study of cytotoxins, many scholars have noticed that traces of phospholipase A will appear in traditional separation methods. 2 (PLA 2 ) (0.1~0.5%, w / w) co-eluted with CTX, while PLA 2 It will have a synergistic effect with CTX, which can significantly enhance the hemolysis of CTX and increa...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/435C07K1/18C07K1/20A61P35/00
Inventor 许云禄
Owner FUJIAN MEDICAL UNIV
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