The choice of residue to be used in the cultivation of a fungal species must take into account the structural and chemical composition of the material in order to be successful in their cultivation. Thus, this study evaluated the physical and chemical characteristics of pineapple and açaí residues and their alterations after the cultivation of two Pleurotus ostreatus strains (474 and 542). The fungi were cultivated in pineapple, açaí, and pineapple + açaí residues, which were added to a mixture of brans, and CaCO3 in the proportion of 78:20:2 w/w/w. The physical and chemical parameters and proximate composition of the residues, and the initial and spent substrates were determined, in addition to being analyzed using scanning electron microscopy and X-ray diffraction. In general, among the residues tested, the pineapple substrate showed more interesting nutritional characteristics for fungiculture, as well as structural conformation and greater availability of nutrients. The açaí substrate was not suitable for fungal cultivation due to its structural characteristics and high tannin content. In this sense, knowing the characteristics of the material used as a support for mushroom growth is of paramount importance in order to achieve successful cultivation.