Isolation, structure elucidation, and in silico Topoisomerase II inhibition of medicarpins isolated from Alfalfa ( Medicago sativa )

(alfalfa) extract led to the isolation of sixteen compounds. Their structures were elucidated using NMR and comparison with reported data. Structural analysis of medicarpin, 10-methoxymedicarpin, and 4-methoxymedicarpin isolated from alfalfa revealed that these compounds share key features with reported Topoisomerase II inhibitors, including a bicyclic non-planar framework and two distinct aromatic groups, suggesting their potential as non-intercalative Topoisomerase II inhibitors. A flexible alignment study demonstrated significant structural similarity between medicarpin and podophyllotoxin, a known Topo II inhibitor. 10-Methoxymedicarpin exhibited the highest binding affinity to Topoisomerase II-DNA complex, along with a stable binding mode and favourable interaction dynamics. Molecular dynamics simulations over 100 ns confirmed the stability of 10-methoxymedicarpin-Topo II-DNA complex, with minimal fluctuations. MM-GBSA, ProLIF, PCAT, and FEL studies further validated the compound's strong binding potential and stability. These findings suggest that 10-methoxymedicarpin is a promising candidate for further development as a Topoisomerase II inhibitor and a potential anticancer candidate.