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Process for preparing thrombase

A technology of thrombin and prothrombin, applied in the field of bioengineering, can solve the problems of long production cycle, complicated process, low yield, etc., and achieve the effects of easy automatic control, high extraction efficiency and simple operation

Inactive Publication Date: 2005-03-02
黄耀江
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Chinese patents ZL92112188, ZL93110426 and ZL96103650 respectively propose methods for extracting thrombin from pig blood or cow blood, but these methods all have problems such as complex process flow, long production cycle and low yield

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1 2800 ml of sheep blood was collected, 400 ml of 0.109 mol / L trisodium citrate anticoagulant was added, and centrifuged. Take supernatant anticoagulated plasma 1000ml. The balanced anion chromatography column on the above-mentioned plasma is eluted with an amino acid eluent whose pH is adjusted to be 8 and whose concentration is 0.2mol / L to obtain 883.2ml of eluent of prothrombin, and add 1mol / L CaCl 2 Solution 76.8ml, so that Ca in the prothrombin solution 2+ The final concentration of prothrombin was 0.08 mol / L, and prothrombin was activated for 3 hours at normal temperature and pressure to obtain 960 ml of thrombin solution, which was marked as A1. The thrombin solution was concentrated by an ultrafiltration machine about 5 times to obtain 190 ml of thrombin concentrate. Freeze-dry the above-mentioned thrombin concentrate to obtain white powdery thrombin finished product.

Embodiment 2

[0023] Example 2 Collect 4300ml of pig blood, add 500ml of 0.109mol / L trisodium citrate anticoagulant, and centrifuge. Take supernatant anticoagulated plasma 1000ml. The balanced anion chromatography column on the above plasma is eluted with trisodium citrate eluent whose pH is adjusted to be 7 and whose concentration is 0.1mol / L to obtain 1590ml of eluent of prothrombin . The eluate was desalted by ultrafiltration to obtain 1832.1 ml of prothrombin solution. Add 1mol / L CaCl 2 Solution 137.9ml, so that Ca in the prothrombin solution 2+ The final concentration of prothrombin was 0.07 mol / L, and prothrombin was activated for 3 hours at normal temperature and pressure to obtain 1970 ml of thrombin solution, which was marked as A2. The thrombin solution was concentrated by an ultrafiltration machine about 9 times to obtain 200 ml of thrombin concentrate. Freeze-dry the above-mentioned thrombin concentrate to obtain white powdery thrombin finished product.

Embodiment 3

[0024] Example 3 3200 ml of bovine blood was collected, 400 ml of 0.125 mol / L potassium oxalate solution anticoagulant was added, and centrifuged. Take supernatant anticoagulated plasma 1500ml. The balanced anion chromatographic column on the above-mentioned plasma is used as an eluent composed of trisodium citrate as a buffer, amino acid as a protective agent, and sodium chloride as an ion enhancer, and the pH is adjusted to 7. The chromatographic column was eluted to obtain 1146 ml of eluate of prothrombin. The eluent was desalted by ultrafiltration to obtain 1440 ml of prothrombin solution. Add 1mol / L CaCl 2 Solution 160ml, so that Ca in the prothrombin solution 2+ The final concentration of prothrombin was 0.1 mol / L, and prothrombin was activated for 3 hours at normal temperature and pressure to obtain 1600 ml of thrombin solution, which was marked as A3. The thrombin solution was concentrated by an ultrafiltration machine about 8 times to obtain 200 ml of thrombin con...

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Abstract

The present invention provides process of extracting thrombase from animal blood or human blood. The process includes adsorbing thrombin from plasma with chromatographic medium, eluting the chromatographic column, activating thrombin to obtain thrombase solution, ultrafiltering to concentrate and finally freeze drying to obtain the thrombase product. The process is one normal temperature process, environment friendly, short in production period, high in thrombase extracting rate of 50000 IU / L, simple in operation, and easy in industrial realization.

Description

technical field [0001] The invention relates to the technical field of bioengineering, in particular to a method for preparing thrombin from animal blood. technical background [0002] At present, prothrombin is mainly prepared from animal plasma and human plasma at home and abroad, and then activated by activators to become thrombin. It catalyzes the cleavage of fibrinopeptides A and B in fibrinogen, transforming it into an insoluble fibrin clot. Thrombin is widely used clinically. It is often applied locally as dry powder or solution on wounds and surgical sites to control blood leakage from capillaries. It is mostly used for bone bleeding, bleeding during tonsillectomy and tooth extraction. Sometimes it can also be taken orally for gastric and duodenal bleeding. Thrombin has a good local hemostatic effect and has no side effects. At present, the scope of application of thrombin is expanding day by day, from simple topical application to hemostasis of bleeding in surger...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/74
Inventor 王永成黄耀江裴靖荆强
Owner 黄耀江
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