Plant material and methanolic extract preparation

Melilotus indicus were collected from Shusha in El-Minia Governorate, Egypt. A botanist, Prof. Magdy Hussein Abd El-Twab, identified the collected plants by using the artificial keys from the taxonomy books and through comparisons with herbarium specimens. Plants preserved in the herbarium of Botany and Microbiology Department, Faculty of Science, Minia University (MIN) under the number AM4044.

The aerial parts of the plant were dried and thoroughly grounded into powder. The powdered form (80 g) was extracted using methanol (200 ml) for 72 h by a soxhlet apparatus. Then filtration was performed using Whatman filter paper no. 1 and the extract was evaporated in a rotar vapour at 40 °C to obtain the dried form. The dried powder was transferred to sterile screw caps and stored at − 20 °C. At use, the frozen dried extract was dissolved in dimethyl sulfoxide (DMSO; 100 mg/ml).