The agronomic use of mushroom post-harvest substrates (SPCHs) in horticultural seedbeds could be an interesting alternative for the reuse of these wastes in line with the European circular economy strategy. This work evaluates the potential use of four treatments with different SPCHs, mushroom (-Ch), mushroom (-St), mushroom compost (-CO), and a mixture (SPCH-Ch and SPCH-St) as substrates for lettuce and chili pepper seed germination. The trial was carried out in a germination chamber using commercial compost as a control treatment. The evaluation was based on its chemical (salinity, N and C content), physical (bulk and real density, porosity and water retention) and plant effect (germination and biomass) characteristics. Of the chemical properties studied, the high salinity in SPCH-Ch and SPCH-CO was a limiting factor for the development of the horticultural species evaluated (electrical conductivity 1:2.5; p/v; ~11 dS m^-1), and low germination percentages were observed. Regarding physical properties, porosity and water retention, the SPCH-CO, SPCH-St and mixture treatments presented some values outside the optimal range established for germination substrates. In the case of SPCH-St, its high C/N ratio could be a limitation for supplying N to the crop. In relation to biomass production (aerial and root) of lettuce and chili pepper, all the treatments evaluated obtained similar values to the control treatment. The mixed treatment presented the highest biomass values, significantly higher in the lettuce crop. In general, the mixed treatment proved to be the best alternative for use in the seedbed.