The Orchidaceae is one of the best represented plant families in Venezuela with nearly 1506 species and 213 genera. Detailed floristics and inventories, however, are needed for different unexplored and underexplored areas of the national parks of Venezuela. The current study was conducted to develop a comprehensive inventory of Orchidaceae in Guatopo National Park (GNP), to assess diversity and distribution, to enhance our knowledge of the local flora, and to determine their conservation priorities. A total of 179 species representing 87 genera of Orchidaceae are reported here. The genera with most species are Epidendrum (18), Pleurothallis (10), Stelis (8), Maxillaria (7), Scaphyglottis (7) Dichaea (5), Maxillariella (5), Prosthechea (5), and Xylobium (5). About 13.40% of the taxa reported in the park are under some categories of the IUCN Red List for threatened species. GNP is one of the protected areas of the north-central region of Venezuela that requires further studies from the perspective of its great biodiversity and in particular to assess the status of existing orchid populations that are under several local anthropogenic threats.