Olive ripening and cultivar effects on carbon stable isotope compositions of n-alkanes and fatty acids in extra-virgin olive oils

The valuable market of extra virgin olive oil (EVOO) requires authentication of geographic origin and variety. Stable isotope analysis of bulk olive oils has been used previously for authentication purposes (Iacumin et al. 2009, Camin et al. 2010a, Camin et al. 2010b, Portarena et al., 2014, Chiocchini et al, 2016). The bulk isotope composition of olive oil represents an average of a relatively large number of isotopically heterogeneous compounds, whose relative proportions may vary among different cultivars and environments (Mihailova et al, 2015). Therefore, variations in isotopic ratios at the bulk level may be influenced by a change in stomatal conductance and/or by a modification in the biochemical composition of oil. Establishing the links between biochemical and physiological responses of different cultivars of olive trees (Olea europaea L.) under various climatic conditions is a important issue for identifying the authenticity and geographical location of olive oils (Portarena et al., 2015). For example, comparing the EVOO's isotopic compositions among varieties with different metabolism (e.g., oil accumulation pattern) may mask the isotopic differences resulting from climate gradients among oil producing regions. Studying isotopic compositions of individual components in EVOO, however, is more suited to traceability studies, particularly when investigating those organic compounds (sterols, aliphatic alcohols, glycerol, fatty acids, n-alkanes) with direct link to environmental factors or sensitive to the biosynthesis stages (Portarena & Brugnoli 2016). To our knowledge, no published studies on combined carbon isotope ratios (?13C) analyses of fatty acids (FAs) and n-alkanes in olive oils are currently available. In this project, bulk oil ?13C as well as n-alkane (n-C29, n-C31, n-C33) and FAs (16:0, 18:0, 18:1, 18:2) ?13C values have been determined to investigate the link between 13C/12C composition of the components and environmental conditions during oil accumulation. In particular, 80 monovarietal EVOO samples were analyzed, representing 4 cultivars in a comparative field and 5 ripening stages. The ?13C values of n-C33 alkane varied among the studied varieties and showed phenological sensitivity, appearing as a useful tool for studying inter-varietal differences in olives. The ?13C of FAs were mainly influenced by ripening stage, without significant effects of the cultivar. Therefore, the environmental conditions mostly determined the ?13C value of FAs during oil accumulation. Thus, combining the bulk ?13C and those of specific compounds, such as n-alkanes and FAs, could provide a better knowledge of EVOO biosynthesis as well as a powerful tool to detect possible adulterations characterizing both the variety and the geographical origin EVOOs.