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Use of cereour bacillus in preparing thrombus treating medicine

A technology for Bacillus cereus and thrombosis diseases, applied in the field of biomedicine, can solve the problems of large side effects, high price, strong toxicity and the like

Inactive Publication Date: 2006-11-08
CHENGDU DIAO JIU HONG PHARMACEUTICAL FACTORY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Thrombolytic drugs used in clinical treatment today, such as streptokinase (Streptokinase, SK), urokinase (Urokinase, UK), recombinant tissue-type plasminogen activator (Recombinant Tissue-type Plasminogen Activator, rt-PA), etc. Certain defects: strong toxicity, large side effects, short half-life in vivo, high price, etc.
So far, there is no relevant report on the use of Bacillus cereus in the preparation of drugs for the treatment of thrombotic diseases, and there is no related report on the use of the fibrinolytic enzyme produced by Bacillus cereus in the preparation of drugs for the treatment of thrombotic diseases

Method used

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  • Use of cereour bacillus in preparing thrombus treating medicine
  • Use of cereour bacillus in preparing thrombus treating medicine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] The preparation of embodiment 1 seed liquid

[0052] After opening the batch of working seed strains, draw 2 to 3 NA medium plates, pick 8 to 10 typical colonies from the plates after culture, and store them on the slant of NA test tubes after culture respectively, and pass the inspection of morphology, purity and production capacity Later as seeds for production.

[0053] Pick a ring of culture from the slant of the checked strain and put it in 20ml NB medium, cultivate it at 37°C and 200rpm for 20h (as the primary seed), then inoculate it in 400ml NB medium at 37°C at 5% inoculum size, Cultivate at 180-220rpm for 18h as seed liquid for fermentation.

Embodiment 2

[0054] Example 2 Preparation of Thrombolytic Enzyme by Liquid Fermentation

[0055] The composition of the fermentation medium is (g / L): sucrose 5g / L, soybean peptone 12g / L, Na 2 HPO 4 12H 2 O 0.50g / L, NaH 2 PO 4 2H 2 O 2g / L, CaCl 2 0.08g / L, MgSO 4 ·7H 2 O 1.0g / L, pH7.5±1.0, sterilized at 121°C for 20min. Sucrose can be replaced by glucose, glycerin, soluble starch, molasses, etc. Soybean peptone can be replaced by corn steep liquor, yeast extract, beef extract, peptone (plant, animal), soybean milk, etc.

[0056] 250ml of the prepared seed solution was added to 1000ml of fermentation medium according to the inoculum size of 12%. The fermentation culture is based on 37°C, 180rpm cultivation for 50h. Centrifuge at 4000rpm for 30min to remove the bacteria, the supernatant is concentrated to 1 / 10 of the original volume with a filter membrane with a cut-off molecular weight of 8000Da, and the auxiliary materials stearate and gelatin are added to make the solid content ...

Embodiment 3

[0064] Embodiment 3 solid-state fermentation prepares thrombolytic enzyme

[0065] Nutrient solution (g / L): glycerin 100g / L, peptone 150g / L, beef extract 80g / L, Na 2 HPO 4 12H 2 O 18g / L, NaH 2 PO 4 2H 2 O 15g / L, CaCl 2 10g / L, MgSO 4 ·7H 2 O 5g / L, pH7.5±1.0.

[0066] Take 1000g of fresh bean dregs, spray 100ml of nutrient solution, cook at 100°C for 30min, and cool; add the prepared seed solution into the fermentation medium according to the inoculation amount of 20%. At 40° C., humidity ≥ 80%, after culturing for 40 hours, dry to obtain the finished product of wax kinase.

[0067] Purification of wax kinase:

[0068] The finished product of wax kinase was leached with 10 times the volume of normal saline, the leaching liquid was sonicated for 20 min, filtered, and the supernatant was collected. The supernatant was prepared into a solution containing 20 mM PB (pH 8.0) at a final concentration. Equilibrate DEAE Sepharose with 20mM pH8.0 PB buffer solution TM Fast ...

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Abstract

The present invention relates to a kind of streptokinase, and is especially one kind of Bacillus cereus produced fibrinolysin, or ceryl kinase. The ceryl kinase has HPLC fingerprint with peak within 7-9 min in the conditions of chromatographic column Waters Protein -PakTM 60 7.8*300 mm, mobile phase 0.2mol / L NaH2PO4-CH3OH(95:5), flow rate 1ml / min and detecting wavelength 220 nm. The present invention also provides the preparation process of the ceryl kinase, and the use of the Bacillus cereus and the ceryl kinase in preparing thrombus treating medicine. The ceryl kinase of the present invention, the rat carotid artery thrombosis experiment shows, possesses obvious thrombosis resisting effect, so that the present invention provides one new option for treatment of thrombus diseases.

Description

technical field [0001] The invention relates to the use of bacillus cereus in the preparation of medicines for treating thrombotic diseases, in particular to the use of fibrinolytic protease produced by bacillus cereus in the preparation of medicines for treating thrombotic diseases, which belongs to the field of biomedicine. Background technique [0002] Thrombosis disease is one of the main diseases that endanger human health, such as cerebral thrombosis and acute myocardial infarction, the mortality rate has reached more than 40%, and thrombosis disease has gradually become the disease with the highest mortality rate. Thrombolysis is an important means of treating such diseases. Thrombolytic drugs used in clinical treatment today, such as streptokinase (Streptokinase, SK), urokinase (Urokinase, UK), recombinant tissue-type plasminogen activator (Recombinant Tissue-type Plasminogen Activator, rt-PA), etc. Certain defects: strong toxicity, relatively large side effects, sh...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/48A61K35/74A61K35/742A61P7/02A61P9/10C12N9/48
CPCC12N9/52C12N9/54A61K38/00A61K35/742C12Y304/21062A61P7/02A61P9/00A61P9/10A61K38/48C12N1/20
Inventor 陈军魏琪蒋飞赖亚娜何俊蓉
Owner CHENGDU DIAO JIU HONG PHARMACEUTICAL FACTORY
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