Fruit tree cultivation requires rootstocks that are resistant to both biotic and abiotic stresses. The container size and substrate used are essential components in their development. Despite this, there are few studies on the impact of substrates on plant development in citrus trees under nursery conditions. This study aimed to assess the effects of three different ratios of pine bark in the substrate of three developing citrus rootstocks in a protected environment (greenhouse). The study conducted at the Cazones Nursery in Cazones de Herrera, Veracruz, Mexico, hypothesized that an increase in bark proportion would lead to a rise in the physical and chemical characteristics of the substrate and the development of the three rootstocks. The study utilized a completely randomized design with a factorial arrangement (A × B). Factor A (rootstock) had three levels: Citrus aurantium L. (Sour Orange), C. volkameriana Pasq. (Volkamer Lemon), and C. sinensis L. × Poncirus trifoliata L. (Citrage C-35). Factor B (substrate) had four levels (0, 10, 20, and 30 % pine bark), resulting in 12 treatments with 20 repetitions each. The physical and chemical characteristics of the substrates were determined, and the plant height and stem diameter were measured. Pine bark positively affected the apparent and real densities, total porosity, electrical conductivity, and cation exchange capacity. The growth dynamics of the three rootstocks were greater during the second and third months after grafting. When grown in substrates with a total porosity of 46–54 %, Volkamer Lemon, Citrage C-35, and Sour Orange rootstocks reached a plant height of 124.1, 110.5, and 84.5 cm, respectively; the stem diameter reached 6.9 mm. Porosity and cation exchange capacity increased when pine bark was added to the substrates. By evaluating the substrates and managing them proportionally, it is possible to obtain plants suitable for grafting (with 5 to 6 mm of stem) within four months after transplanting. This results in less time spent in the nursery and reduced costs.