Abstract

PRODUCTION AND CHARACTERIZATION OF FIBRINOLYTIC ENZYME FROM ASPERGILLUS NIGER.

Pallavi K. Aradhye*, Meera D. Chavan

ABSTRACT

Background: Cardiovascular diseases are the leading cause of death for men and women. Heart attack and stroke account for more deaths than all cancers and injuries combined. To sort this problem microbial fibrinolytic enzyme may found the efficient therapeutic treatment. Objective: To isolate and study fibrinolytic enzyme from fungal source. Method: Aspergillus niger was cultivated in milk sabouraud’s broth for the production of proteases. Extraction of enzyme was carried out by filtration. For partial purification acetone precipitation and dialysis preferred. Protease activity was measured with casein and fibrin substrate. Protein concentration was determined by folin Lowry method. Effect of various environmental factors such as temperature, pH and metal ions on enzyme activity was tested. Electrophoresis was performed for molecular weight determination. Fibrinolytic activity studied on fibrin agar plate and blood clot. Result: Maximum enzyme production was achieved in 10 days of incubation at 300C ± 20C; with an initial pH of 5.6 the protein fraction precipitated with 50% acetone had the fibrinolytic activity 64.64U/mg protein. This partial purification was to 4.48 folds. The optimum activity of the enzyme was reached at 400C and pH, 7.0 with 0.5 % of substrate casein concentration. The enzyme activity was enhanced by Ca2+ and Mg2+ but was inhibited by Co, Cu, Zn, and Fe at 1mM concentration. The enzyme showed band at 31 kDa after SDS-PAGE. The enzyme showed clear zone on 1.5% fibrin plate when incubated at 370C for 18 h. Human blood clot lysis checked after 19 h of incubation at 370C. Conclusion: These results indicate production of fibrinolytic enzyme was found to be potent fibrinolytic agent.

Keywords: Fibrinolytic enzyme, Aspergillus niger, Fibrin plate, Blood clot lysis.