Spirostanol saponins and esculin from Rusci rhizoma reduce the thrombin-induced hyperpermeability of endothelial cells

Abstract Rusci rhizoma extracts are traditionally used against chronic venous disorders (CVD). To determine the effect of its secondary plant metabolites on the endothelium, phenolic compounds and saponins from Butcher’s broom were isolated from a methanolic extract, and their activity on the thrombin-induced hyperpermeability of human microvascular endothelial cells (HMEC-1) was investigated in vitro . In addition to the six known spirostanol saponins deglucoruscin ( 5 ), 22-O-methyl-deglucoruscoside ( 6 ), deglucoruscoside ( 7 ), ruscin ( 8 ), ruscogenin-1-O-(α- l -rhamnopyranosyl-(1 → 2)-β- d -galactopyranoside ( 9 ) and 1-O-sulpho-ruscogenin ( 10 ), three new spirostanol derivatives were isolated and identified: 3′-O-acetyl-4′-O-sulphodeglucoruscin ( 1 ), 4′-O-(2-hydroxy-3-methylpentanoyl)-deglucoruscin ( 2 ) and 4′-O-acetyl-deglucoruscin ( 3 ). Furthermore, the coumarin esculin ( 4 ), which is also prominently present in other medicinal plants used in the treatment of CVD, was isolated for the first time from Rusci rhizoma . Five of the isolated steroid derivatives ( 2 , 5 , 8 , 9 and 10 ) and esculin ( 4 ) were tested for their ability to reduce the thrombin-induced hyperpermeability of endothelial cells in vitro , and the results were compared to those of the aglycone neoruscogenin ( 11 ) . The latter compound showed a slight but concentration-dependent reduction in hyperpermeability to 71.8% at 100 μM. The highest activities were observed for the spirostanol saponins 5 and 8 and for esculin ( 4 ) at 10 μM, and these compounds resulted in a reduction of the thrombin-induced hyperpermeability to 41.9%, 42.6% and 53.3%, respectively. For 2, 5 and 8, the highest concentration tested (100 μM) resulted in a drastic increase of the thrombin effect. The effect of esculin observed at a concentration of 10 μM was diminished at 100 μM. These in vitro data provide insight into the pharmacological mechanism by which the genuine spirostanol saponins and esculin can contribute to the efficacy of Butcher’s broom against chronic venous disorders.