In this study, calli of Medicago sativa L. cv. Elçi (alfalfa Elçi) were inoculated in cell suspension culture and analyzed for aggregate assay, cell viability test, total phenolic content assay, DPPH free radical scavenging activity and formononetin assay by means of High-Performance Liquid Chromatography (HPLC). Hypocotyl, cotyledon and apical meristem explants were taken from 15-day-old aseptic seedlings and germinated in MS medium. 10 g calli were grown for each explant and then transferred into cell suspension culture. The highest cell viability rate, which was 75%, and the highest DPPH free radical scavenging activity with 51.36% was measured in 1000 mL cell suspension culture, while the highest total phenolic content, i.e. 40.2 mg/g, was quantified in 250 mL cell suspension culture. In accordance with the findings of the study, the production of formononetin was higher in the calli derived from cell suspension cultures than in herb samples of M. sativa. Moreover, in 1000 mL cell suspension culture, 4.99 mg/g of formononetin concentration was quantified, which scored the highest. In large-scale cell suspension cultures of M. sativa, it was possible to increase the production of formononetin production. Hence, due to its medicinal significance, a method has been tested to obtain higher amounts of this compound.
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