New sesquiterpene lactones from arnica tincture prepared from fresh flowerheads of Arnica montana

Investigation of an ethanolic extract prepared from fresh Arnica montana flowers afforded three new 1,5-trans-guaianolides, of which 11 alpha, 13-dihydro-2-O-tigloylflorilenalin and the respective 2-O-isovaleryl derivative are reported for the first time. Additionally, three new and one known 2 beta-ethoxy-2,3-dihydrohelenalin esters were isolated. GC/MS studies of the extract after a two year storage at 4 degrees C demonstrated that the latter were artefacts that had been formed by addition of ethanol to the cyclopentenone structure of helenalin. Formation of these adducts gave compounds possessing an inhibitory activity comparable to that of 11 alpha,13-dihydrohelenalin derivatives in the NF-kappa B EMSA and the IL-8 ELISA in vitro assays as well as in the in vivo croton oil-induced mouse ear edema test for one adduct, namely 2 beta-ethoxy-6-O-acetyl-2,3-dihydrohelenalin. As expected, 6-O(2-methylbutyryl)- and 6-O-methacryloyl-helenalin exhibited a stronger activity in the NF-kappa B EMSA and IL-8 ELISA. Sesquiterpene lactones seem to be the most important NF-kappa B inhibiting compounds in the Arnica extract. Bioguided fractionation using the luciferase reporter gene assay resulted in the isolation of only moderately active compounds, such as 6-acetoxy-2,2-dimethylchroman-4-one and 10-acetoxy-8,9-epoxythymol isobutyrate.