Evidence of the Direct Reaction Between Ozone and Molecular Components in Epicuticular Waxes of Olive Leaves and Grape Berry Skins

Tropospheric ozone has been found to have detrimental effects on plant life, in part due to the degradation of epicuticular waxes which protect leaves and grape berries from water loss, ultraviolet radiation, pathogens, and insects. A number of studies have explored the morphological changes and broad changes in functional group composition in epicuticular waxes due to ozone exposure. However, little work has been done on direct reactions between molecular components in epicuticular wax and ozone. Our research focuses on investigating the reactions between ozone and two important molecular components of epicuticular waxes—oleanolic acid and ferulic acid. Pure standards of oleanolic acid and ferulic acid were exposed to ozone in PTFE bottles under dry and humid conditions and the products were separated using flash chromatography. Pure products were analyzed using HPLC-QToF-MS and a variety of 1D and 2D NMR techniques. Epicuticular waxes extracted from the leaves of a dwarf olive tree (Olea europea) and the skins of red and green table grapes were ozonized in PTFE bottles under humid and dry conditions and then analyzed via HPLC-QToF-MS. The chromatograms and mass spectra for the reacted waxes were compared to those for ozonolysis products of pure standards. We determined that oleanolic acid reacted with ozone to form a lactone product in waxes from both Olea europa leaves and grape berry skins. In addition, we found a new potential cross-reaction product resulting from the reaction between oleanolic acid and ferulic acid in the presence of ozone in the reacted leaf waxes from the dwarf olive tree. We are currently working to identify the cross-product.