Method for fermenting and producing plasmin by use of cordyceps militaris liquid

A technology of liquid fermentation and fibrinolytic enzymes, applied in the direction of using fungi, methods based on microorganisms, biochemical equipment and methods, etc., can solve problems such as no clear conclusions, achieve improved liquid fermentation production, low cost of raw materials, Fermentation produces good results

Inactive Publication Date: 2012-07-25
INST OF AGRI RESOURCES & ENVIRONMENT SHANDONG ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is still no clear conclusion about the mechanism of action of expansin. There are no reports at home and abroad on the application of expansin in the liquid fermentation of the edible and medicinal fungus Cordyceps militaris for the production of plasmin and other secondary metabolites.

Method used

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  • Method for fermenting and producing plasmin by use of cordyceps militaris liquid

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0036] The preparation steps of the expansin solution described in the examples are as follows:

[0037] Broad bean (Vicia fabaL.; purchased from Jinan Maofeng Seedling Co., Ltd.) or cucumber (Cucumis sativus L. CV. Jinnian No.6; purchased from Jinan Weili Seed Industry Co., Ltd.) seeds were treated with 0.15 wt% HgCl 2 Disinfect for 6 minutes, rinse with running water for 6 hours, plant in wet vermiculite, and culture in dark at 27°C for 4 days. Cut the epicotyl of the seedlings to 4-5cm, that is, the growth area is about 100g, pre-cool in the refrigerator at -20°C for 1 hour, add the homogenization buffer pre-cooled to 4°C, and after high-speed homogenization, filter with 70μm nylon mesh, and filter the residue through Wash with homogenization buffer, then add the filter residue to the homogenization buffer, stand at room temperature for 2 hours to obtain a standing solution; add the extract to the standing solution, extract at 4°C for 24 hours, filter, and the filtrate is 0...

Embodiment 1

[0051] A liquid fermentation method utilizing expansin to increase the yield of the plasmin component of Cordyceps militaris, comprising the steps of:

[0052] (1) Cordyceps militaris (Cordyceps militaris) strain, the strain number is CGMCC No.5.700, was inoculated into PD liquid fermentation medium, and activated culture was carried out. Cultivate for 48 hours, then transfer to the seed fermentation medium with an inoculation amount of 15% by volume, and carry out seed cultivation. The seed cultivation conditions are: temperature 25° C., shaker cultivation for 48 hours, to obtain seed liquid;

[0053] (2) Inoculate the seed solution prepared in step (1) in a liquid fermentation medium at a volume ratio of 15%, and culture it for 24 hours in liquid fermentation at a temperature of 25° C., and then add the expansin solution to make the concentration of the expansin 1.5mg / mL, then continue to culture for 144h, centrifuge for 10min under the condition of 12000r / min, remove the my...

Embodiment 2

[0068] The liquid fermentation method as described in Example 1, the difference is that in step (2), at a temperature of 25° C., the expansin solution is added to make the concentration of the expansin 2.0 mg / mL, and then the culture is continued for 144 hours.

[0069] After detection and calculation, the activity of the fibrinolytic enzyme component per milliliter of Cordyceps militaris fermentation liquid is 93.27U, that is, the activity of the fibrinolytic enzyme component per liter of Cordyceps militaris fermentation liquid is 93270U; the results are as follows figure 1 shown.

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Abstract

The invention relates to a method for fermenting and producing plasmin by use of cordyceps militaris liquid. The method comprises the following steps of: (1) inoculating the cordyceps militaris strain into the PD liquid fermentation medium to perform activation culture, and transferring to a seed fermentation medium to perform seed culture to obtain seed liquid; (2) inoculating the seed liquid into a liquid fermentation medium according to a volume ratio of 5-15%, and performing liquid fermentation culture for 20-30 hours; adding an expansion protein solution, and continuing culture for 120-168 hours; and separating and removing cordyceps militaris mycelium to obtain plasmin fermentation supernate; and (3) extracting cordyceps militaris extracellular plasmin from the plasmin fermentation supernate to obtain the cordyceps militaris plasmin. According to the invention, the plant expansion protein and the optimized fermentation method are applied to the liquid fermentation production of cordyceps militaris plasmin, and the liquid fermentation output of the cordyceps militaris plasmin is remarkably increased to 93270U per liter of fermentation broth, which is over 2.69 times that of the traditional fermentation production method; and the method has perfect industrial application prospect.

Description

technical field [0001] The invention relates to a method for producing fibrinolytic enzyme by liquid fermentation of Cordyceps militaris, belonging to the technical field of biological fermentation engineering. Background technique [0002] Thromboembolic disease is currently one of the diseases that endanger human health and cause the highest mortality rate, including acute myocardial infarction, ischemic heart disease, cardiovascular disease, heart valve disease, peripheral vascular disease, arrhythmia, hypertension, cerebral embolism , Pulmonary embolism, etc. increasingly threatening human health. According to a study published in the American Heart Association's Circulation: Cardiovascular Quality and Outcomes, the annual incidence of cardiovascular disease worldwide is projected to increase by 50% between 2010 and 2030 alone due to aging and population growth. %. At present, the most effective and reliable means of treating thromboembolic diseases is thrombolytic th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/68C12R1/645
CPCC12N1/14C12N9/48C12N9/6435C12N9/14C12N9/00C12R1/645
Inventor 姚强宫志远单洪涛韩建东王琦高能孙涛任鹏飞万鲁长任海霞李瑾
Owner INST OF AGRI RESOURCES & ENVIRONMENT SHANDONG ACADEMY OF AGRI SCI
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