Volatile constituents of Trifolium pratense spp. nivale quantified at different growth stages, and evaluation of their antimicrobial activity.

The composition of the volatile fraction of Trifolium pratense L. ssp. nivale (Sieber) Asch. et Gr. from the southwestern Alps was investigated. Fresh aerial parts were collected in the summer at three different growth stages, namely vegetative, flowering and fruiting. The oils obtained by steam-distillation accounted for 0.006 to 0.011% of the fresh plant material and their composition was determined by GC/FID and GC/MS. Several classes of compounds were found, with a predominance of alcohols in all phases, followed by aldehydes, hydrocarbons, terpenes, phenolics, ketones, acids and esters. The oil composition varied both in quantity and quality; the most abundant compounds were oct-1-en-3-ol and phenylacetaldehyde in the vegetative and flowering phases, and phenylacetaldehyde and 2-phenylethanol in the fruiting phase. The essential oils obtained were tested for activity against two major bee pests, i.e. Paenibacillus larvae and Melissococcus plutonius, and against a reference bacterial species, Bacillus subtilis.