Secondary metabolite extraction

The fungi were cultivated on PDB by inoculating selected endophyte cultures in 250 mL Erlenmeyer flask containing 100 mL of the medium. The flask was incubated at 28 °C for 2 week with periodical shaking at 150 rpm. After the incubation period, the fermentation broth of the fungus was homogenized by adding 10 % methanol to it. Metabolite was extracted by solvent extraction procedure using ethyl acetate and methanol as organic solvents. To the filtrate equal volume of solvents were added, mixed well for 10 min and kept for 5 min till the two clear immiscible layers formed. The upper layer of solvent containing the extracted compounds was separated using separating funnel. Solvent was evaporated and the resultant compound was dried in rotator vacuum evaporator to yield the crude metabolite (Bhardwaj et al. 2015). The crude extract was then dissolved in Dimethyl sulphoxide at 1 mg/mL of concentration and kept at 4 °C.