Thiophenes and triterpenes from Eclipta prostrata and their inhibitory effects on steroid 5α-reductase type II

Four new thiophenes ( 1 - 4 ) and one new nortriterpenoid saponin ( 5 ), alongside 31 known compounds, were isolated and structurally elucidated from the ethyl acetate extract of Eclipta prostrata . In bioassay, 3-oxo-16α-hydroxy-olean-12-en-28-oic acid ( 26 ) and echinocystic acid ( 19 ) exhibited potent inhibitory activity against steroid 5α-reductase type II with outstanding IC 50 values of 7.54 ± 0.26 and 15.87 ± 2.30 μ M, respectively. Enzyme kinetic analyses revealed that compounds 26 and 19 were both non-competitive inhibitors of steroid 5α-reductase type II. Molecular docking studies provided supportive evidence that compounds 26 and 19 did not directly compete with testosterone, but might induce conformational changes in the active site of steroid 5α-reductase type II, thereby inhibiting the production of dihydrotestosterone. In addition, molecular dynamics simulations supported the formation of stable complexes between steroid 5α-reductase type II and compound 26 or 19 via strong intermolecular interactions. Synergistic assays showed that echinocystic acid ( 19 ) and wedelolactone ( 32 ) exhibited the strongest synergy (ZIP = 27.597), and the optimal ratio was 1:4 to produce the lowest combination index value.