Helleborus species (family Ranunculaceae) are evergreen, rhizomatous plants with scientifically demonstrated biological/pharmacological activities [1]. Nonetheless, the essential oils of the genus are general poorly phytochemically and pharmacologically investigated. Helleborus odorus Waldst. & Kit. ex Willd. (‘fragrant hellebore’ or ‘kukurek’ in Serbian) is a highly toxic plant species growing on hillsides and in forests of the submediterranean region. Up to now, steroid-related compounds (saponins, ecdysteroids, bufadienolides), fatty acids and other lipids, and sugars of H. odorus were investigated. However, to the best of our knowledge, there are no previous studies of the essential oil of this species. Analyses by GC and GC/MS of four essential-oil samples obtained from dry aerial parts and roots of H. odorus allowed the identification of 229 components, comprising 88.8-92.2% of the total oil composition. The major identified volatile compounds were 1-pentacosene (0.0-52.8%), tricosane (0.0-15.1%), linoleic acid (0.0-11.8%), trans-phytol (0.0-12.4%), hexadecanoic acid (3.7-16.8%), (2E,4E)-decadienal (tr-13.8%), linalool (0.5-6.0%) and hexanal (0.0-7.4%). In general, there were qualitative and quantitative variations noted in the compositions between the H. odorus essential-oil samples from different locations. The most discernable differences included a change in the content of the major constituent (1-pentacosene). These differences motivated us to explore (by multivariate statistical analysis (MVA)) the possible existence of several chemotypes of this species using essential-oil compositional data. Agglomerative hierarchical clustering and principal component analysis of the chemical data on the volatiles of the herein studied and additional 37 oil samples of Ranunculaceae showed a close relationship of Helleborus with other Ranunculaceae genera and the existence of only one chemotype of this species in the Serbian flora
