This study evaluates the effects of novel peat-free organic substrates and a novel liquid fertilizer on the biometric features and macronutrient allocation of Quercus robur and Fagus sylvatica seedlings with the view to compare biomass and nutrient allocation of plant organs in seedlings cultivated on peat substrates against those grown on novel peat-free substrates and fertilizer. The experimental setup involved four substrate variants, including the control (R20, R21, R22 and C). The novel substrates and fertilizer were designed and formulated by the University of Agriculture in Kraków, Poland (UAK). Fertilization used in the state forest nurseries was represented as SR20, SR21, and SR22, while the novel fertilizer of UAK was represented as UR20, UR21, and UR22; meanwhile, SC and UC represented the control substrates (peat) in both cases, respectively. The experiment was laid in a 2x2x4 experimental design using five seedlings per treatment. Seedlings were assessed for height, collar diameter, and roots, shoots, and leaves biomass. The allocation patterns highlighted the variability of nutrient allocation within the plants, with more nutrients allocated to the root system. Interestingly, treatment UR22 yielded the highest mean root values, root biomass, and virtually all macroelement allocation. Overall, peat-free substrates with novel liquid fertilizer demonstrated competitive traits supporting seedling growth, with significant differences observed in the allocation of nutrients like nitrogen, phosphorus, and potassium. Based on the study results, peat-free substrates with the recommended dosage of novel fertilizer improved seedlings' quality, enhancing the growth and nutrient allocation in different organs of tree seedlings.