The Péclet correction is often used to predict leaf evaporative enrichment and requires an estimate of effective path length (L). Studies to estimate L in conifer needles have produced unexpected patterns based on Péclet theory and leaf anatomy. We exposed seedlings of six conifer species to different vapour pressure deficits (VPD) in controlled climate chambers to produce steady-state leaf water enrichment (in 18O). We measured leaf gas exchange, stable oxygen isotopic composition (δ18O) of input and plant waters as well as leaf anatomical characteristics. Variation in bulk needle water δ18O was strongly related to VPD. Conifer needles had large amounts of water within the vascular strand that was potentially unenriched (up to 40%). Both standard Craig-Gordon and Péclet models failed to accurately predict conifer leaf water δ18O without taking into consideration the unenriched water in the vascular strand and variable L. Although L was linearly related to mesophyll thickness, large within-species variation prevented the development of generalizations that could allow a broader use of the Péclet effect in predictive models. Our results point to the importance of within needle water pools and isolating mechanisms that need further investigation in order to integrate Péclet corrections with 'two compartment' leaf water concepts. This study measured the variation in the stable oxygen isotopic composition of needle water for six conifer species. Although the Péclet correction of leaf water evaporative enrichment has been used extensively in predictive models (many of which have been published in PCThe enigma of effective path length for 18 O enrichment in leaf water of conifers=10.1111/pce.12568=amp;E), conifer needles can sometime produce results that are odd in reference to Péclet theory or leaf anatomy. Due to the importance of conifers for long-lived tree ring isotope records and their massive ecosystem effects on biogeochemical cycles, this study could impact diverse research fields that require accurate estimates of conifer leaf water evaporative enrichment.