Building a Barrier: The Influence of Different Wax Fractions on the Water Transpiration Barrier of Leaf Cuticles

Waxes are critical in limiting non-stomatal water loss in higher terrestrial plants by making up the limiting barrier for water diffusion across cuticles. Using a differential extraction protocol, we investigated the influence of various wax fractions on the cuticular transpiration barrier. Triterpenoids (TRPs) and very long-chain aliphatics (VLCAs) were selectively extracted from isolated adaxial leaf cuticles using methanol (MeOH) followed by chloroform (TCM). The water permeabilities of the native and the solvent-treated cuticles were measured gravimetrically. Seven plant species (Camellia sinensis, Ficus elastica, Hedera helix, Ilex aquifolium, Nerium oleander, Vinca minor, and Zamioculcas zamiifolia) with highly varying wax compositions ranging from nearly pure VLCA- to TRP-dominated waxes were selected. After TRP removal with MeOH, water permeability did not or only slightly increase. The subsequent VLCA extraction with TCM led to increases in cuticular water permeabilities by up to two orders of magnitude. These effects were consistent across all species investigated, providing direct evidence that the cuticular transpiration barrier is mainly composed of VLCA. In contrast, TRPs play no or only a minor role in controlling water loss.