This article presents the results of a study on plant diversity at different levels in residual forest stands, located in the historical Favorita Park in Palermo, Italy (established and named in 1799 by King Ferdinand IV of the House of Bourbon). These forest aspects have naturally evolved for over two centuries, under minimal conditions of anthropogenic disturbance (e.g. deforestation, fires, grazing activities, etc.). This is especially true in the area known as “Bosco Niscemi”, spread over about 8.5 hectares, in the centre of the park. Bosco Niscemi is characterized by the widespread presence of old trees, abundant necromass and litter. In this study, four different soil profiles were analysed, and classified as follows: (i) Solimovic Regosol (Arenic); ii) Eutric Arenosol (Chromic); (iii) and (iv) Skeletic Regosol (Ochric). From a phytosociological point of view, four forest communities have been identified, two of which are described as new associations (Viburno tini-Phillyreetum latifoliae ass. nova and Teucrio flavi-Phillyreetum latifoliae ass. nova). The species richness was also found to be noteworthy, with the co-occurrence of several taxa (phanerogams and cryptogams) that are of biogeographic interest or rare in Sicily, including: i) vascular flora (e.g. Viburnum tinus and Arbutus unedo, both very rare throughout the western sector of Sicily); ii) bryophytes [Cryphaea heteromalla (new record of a very rare species in Sicily) as well as Hypnum cupressiforme and Leptodon smithii, also never previously found at such low altitudes]; iii) lichens (e.g. Bacidia rosella, Gyalecta derivata, Ramalina roesleri and Waynea stoechadiana); iv) mushrooms (e.g. Eichleriella leucophaea, only known location in Italy). Based on the scientific documentation produced in this study, these woods can be reported as “old-growth forests” to be included in the “National Network” that has been recently established in Italy (Ministerial Decree of 23 June 2023). This area might serve as an ideal control for urban environmental studies, given its pristine ecological setting.