Erythropalum scandens Bl. is a medicinal woody vegetable found in southern China and parts of Southeast Asia. Studies have shown improper substrate hindered E. scandens seedling growth, causing water accumulation and nutrient deficiency. In pursuit of an ideal growth medium for E. scandens seedlings during the early stages, this study conducted a pot experiment to identify a mixed substrate with optimal water permeability and fertility. In this study, pure Alfisols soil treatment as the control (CK), and two soilless substrates (peat soil and perlite) were combined with Alfisols soil into different volume ratios, in order to better use soil resources from understory space and balance the texture of mixed substrates. The growth, physiological characteristics and nutrient status of 24-month-old E. scandens seedlings were determined after planting in different mixed ratios. The results showed that as the proportion of peat soil increased in the mix, most indexes exhibited an initial increase followed by a decline, while soluble protein content decreased consistently. Conversely, an increasing perlite ratio resulted in a general decline in most growth and physiological indexes. Root growth, biomass accumulation and chlorophyll content, peaked in the 66.67% Alfisols soil + 33.33% perlite (T4) treatment. Notably, T3 (66.67% Alfisols soil + 33.33% peat soil) showcased the best above-ground growth, while T1 (50.00% Alfisols soil + 50.00% peat soil) excelled in element content accumulation. In conclusion, the cultivation substrate should primarily consist of Alfisols soil, constituting at least 50%. The addition of peat soil enhances above-ground growth and nutrients accumulation, while perlite contributes to robust root development. One third of peat soil and a small amount of perlite can be added to the substrate during E. scandens seedling cultivation, and proper fertilization should also be used in order to increase nutrient accumulation in aboveground and underground parts. This research provides valuable insights into maximizing the potential of E. scandens seedlings through precise cultivation methods.