The aim of this study was to establish the possibility of growing Rosa canina L. and Pelargonium zonale L. in tissue culture and suitability of alternative components and their mixtures as a substrate for adaptation of in vitro seedlings of these floral species. In this research, four alternative components (cocoa shell (A), willow bark (B), spent mushroom substrate (C), sawdust (D)) and six of their mixtures were analysed. Klasman Potgrond P (for wild rose) and Balkon – blumenerde (for pelargonium) were analysed as a control substrate. Statistical analysis has revealed significant differences between all the treatments (substrates) and their analysed properties. An average percentage of adapted seedlings in all of the substrates was 54.68% for wild rose, and 99.68% for pelargonium. The largest values of morphological properties for the wild rose were determined on an M2 (30% A + 20% B + 40% C + 10% D) substrate mixture, where the greatest number of shoots, leaf number, fresh overground mass, total fresh mass of seedling and the ratio of fresh and dry overground mass and root were recorded. Chemical composition of wild rose seedlings varied among the treatments, but the highest content of macro and micro elements was determined in the seedlings grown on sawdust (D) and control substrate (E). The largest values of almost all morphological properties of pelargonium were also recorded on substrate mixtures M4 (18% A + 22% B + 40% C + 20% D) and M6 (18% A + 42% B + 20% C + 20% D). Furthermore, while the largest morphological properties of pelargonium seedlings were recorded in the mixtures, the highest content of macro and micro elements determined in the seedlings was on those grown on alternative components, especially in the seedlings grown on a willow bark substrate.