Trimeresurus albolabris defibrase preparation method

A technology of bamboo leaf green and defibrase, which is applied in the directions of medical preparations, hydrolase, and pharmaceutical formulas containing active ingredients.

Inactive Publication Date: 2010-01-27
CHONGQING NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, there is no effective method for extracting and purifying the defibrase from the venom of the white bamboo leaf green snake in my country and the confirmation of its efficacy

Method used

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  • Trimeresurus albolabris defibrase preparation method
  • Trimeresurus albolabris defibrase preparation method
  • Trimeresurus albolabris defibrase preparation method

Examples

Experimental program
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Effect test

Embodiment 1

[0023] (1) Pretreatment of the venom of the green snake:

[0024] The venom was collected from the green snake venom by biting the dish, and the collected venom of the green snake was concentrated and then freeze-dried to obtain the lyophilized powder with crude poison. Take 1 g of the venom crude freeze-dried powder of the green snake venom and dissolve it in 5ml pH8.0, 0.05mol Tris-HCl buffer solution, centrifuge at 3000r / min for 10min with a centrifuge, and take the supernatant for later use;

[0025] (2) DEAE-Sephadex A-25 ion exchange chromatography

[0026] With concentration 0.05mol / L, the Tris-HCl buffer solution of pH8.0 equilibrates SephadexA-25 column, chromatographic column: 2cmx60cm; The supernatant that makes in the step (1) is loaded with the sample, with concentration range 0.6mol / LNaCl1000ml 0.05mol Tris-HCl buffer linear gradient elution, the elution flow rate is 0.25ml / min, (see figure 2 ) The I peak eluent separated by SephadexA-25 column chromatography ...

Embodiment 2

[0032] (1) Pretreatment of the venom of the green snake:

[0033]The venom is collected from the green snake venom by biting the dish, and the collected venom of the green snake is concentrated and then freeze-dried to obtain a lyophilized powder with crude poison. Take 5 g of the venom crude freeze-dried powder of the green snake venom and dissolve it in 5ml pH8.5, 0.06mol Tris-HCl buffer solution, centrifuge at 5000r / min for 12min with a centrifuge, and take the supernatant for later use;

[0034] (2) DEAE-Sephadex A-25 ion exchange chromatography

[0035] Equilibrate the SephadexA-25 column with a Tris-HCl buffer solution with a concentration of 0.06mol / L and pH 7.0, a chromatographic column: 2cm x 60cm; the supernatant obtained in step (1) is loaded with a concentration range of 0.01mol / L L NaCl 1000ml 0.06mol Tris-HCl buffer linear elution, the elution flow rate is 0.30ml / min, freeze-dry the eluate from peak I separated by SephadexA-25 column chromatography, dissolve in ...

Embodiment 3

[0041] (1) Pretreatment of the venom of the green snake:

[0042] The snake venom is collected from the green snake venom by biting the dish, and the collected venom of the green snake is concentrated and then freeze-dried to obtain a lyophilized powder with crude poison. Take 2.5 g of the venom crude freeze-dried powder of the green snake venom and dissolve it in 5ml pH 7.5, 0.04mol Tris-HCl buffer solution, centrifuge at 4500r / min for 8min with a centrifuge, take the supernatant for later use;

[0043] (2) DEAE-Sephadex A-25 ion exchange chromatography

[0044] Equilibrate the SephadexA-25 column with Tris-HCl buffer solution with a concentration of 0.04mol / L, pH 8.0, chromatography column: 2cm x 60cm; the supernatant obtained in step (1) is loaded with a concentration range of 0.6mol / L L NaCl1000ml 0.04mol Tris-HCl buffer linear gradient elution, the elution flow rate is 0.20ml / min, freeze-dry the eluate from peak I separated by SephadexA-25 column chromatography, dissolve...

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Abstract

The invention relates to fibrinolytic enzyme of trimeresurus albolabris venom, and the preparation method and the application thereof in pharmaceutical industry, and belongs to the biomedical field. Trimeresurus albolabris venom fibrinolytic enzyme is metal protease obtained in the venom of the domestic trimeresurus albolabris through separation and purification, has the activity of simultaneously degrading fibrinogen Aalpha chain and fibrinogen Bbeta chain, and is 66300 dalton under the reducing and non-reducing conditions of the apparent molecular weight on polyacrylamide gel electrophoresis. The preparation method comprises the following steps: firstly, pretreatment of trimeresurus albolabris venom; secondly, DEAE-Sephadex A-25 ion-exchange chromatography; thirdly, DEAE-Sephadex G-100 gel chromatography; fourthly, CM-Sephadex C-25 cation-exchange chromatography. The fibrinolytic enzyme of the trimeresurus albolabris venom has higher activity of simultaneously degrading fibrinogen Aalpha chain and fibrinogen Bbeta chain, and can be applied for curing thromboembolic disease.

Description

technical field [0001] The invention provides a preparation method of the defibrase from the venom of the venom of the bamboo leaf bamboo leaf. Background technique [0002] There is a class of enzymes that can directly dissolve fibrin (ogen) in snake venoms such as Viperinae, Viperinae, and Spectacles, referred to as defibrase, which is a kind of enzyme that can dissolve both fibrinogen and fibrin. fibrinolytic enzymes. There is no inactive zymogen form of snake venom defibrase, but there is an active proteolytic enzyme that does not require an activator to activate. Therefore, it can directly act on the fibrin that forms thrombus, dissolve the thrombus quickly, and improve the clinical symptoms of patients; by dissolving plasma fibrinogen, it can reduce blood viscosity, inhibit thrombus formation, and prevent re-infarction. [0003] According to its structural characteristics, snake venom defibrase can be divided into serine proteinase and metalloproteinase. The defibri...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/64A61K38/48A61P7/02
Inventor 余晓东邓敏
Owner CHONGQING NORMAL UNIVERSITY
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