Hydroponics is an excellent alternative approach for the production of seedlings, given the growing demand for fruiting trees for the reforestation or recuperation of degraded natural landscapes. In most cases, however, little is known about the optimal period for the maintenance of the seedling in the hydroponic system. Given this, we decided to investigate the hypothesis that morpho-anatomical and physiological alterations can be used to predict the optimal timing for the transplantation of the seedlings to the soil substrate, thereby guaranteeing the most cost-effective application of the hydroponic system. We selected Hymenaea courbaril L., an important Neotropical fruit tree, as the model for this study. We cultivated H. courbaril seedlings in a static hydroponic system and evaluated morpho-anatomical, physiological, and growth parameters over the course of seedling development (30, 60, 90, 120, 150, and 180 days after transplantation; DAT). We observed an interesting relationship between the increase in the density (SD) and conductance (gsw) of the stomata up to 120 DAT, which reflected higher rates of photosynthesis (A), but also a reduced efficiency in the use of water. In the subsequent intervals, the SD of the plants and the diameter of the radicular xylemic vessels elements (RVE) decreased, in an attempt to increase the efficiency of the use of this resource. We also observed an increase in the thickness of the palisade parenchyma (PP) prior to 120 DAT, which did not reflect a general increase in the thickness of the mesophyll, indicating an adjustment in the thickness of the spongiform parenchyma (SP). We also observed a progressive increase in photosynthetic efficiency up to 120 DAT, based on parameters such as the absorption flux energy per active reaction center (ABS/RC) and the photosynthetic performance index (PIABS), but after this period these indices decreased progressively. However, as the PIABS is an indicator of the plant’s tolerance, its decline was associated with an increase in the dissipation of energy (DI0/RC), which indicates that, after 120 DAT, the plant pots may become a stress factor that limit the growth of H. courbaril seedlings. The results of the present study indicate conclusively that a 120-day period is the optimum for the maintenance of the H. courbaril seedlings in the hydroponic system, and also confirm the hypothesis that the morpho-anatomical and physiological responses observed in the plants can be used to predict the ideal period for the transplantation of the seedlings, contributing to a reduction in production time of the hydroponic system.