This study presents the floristic survey of orchid species occurring in the Serra do Japi, São Paulo, Southeastern Brazil. This region is strategically placed in the transition between interior semi-deciduous mesophytic forests and the Atlantic forest, presenting species from both formations. Although the orchid diversity is high, the region is affected by anthropogenic disturbance. The preservation and the integrated study is an urgent necessity, with social, economic and preservationist reflexes. The observed orchid family includes 125 species distributed among 61 genera. Two new species, Habenaria sp. and Acianthera sp. which were not identified before are described. The most representative genus is Epidendrum, followed by Oncidium and Habenaria. Most of the species occurs as epiphytes, while 40 species are terrestrial, 31 species are rupicolous, two are hemi-epiphytes and only one is a saprophyte. The semi-deciduous mesophytic forest has the highest occurrence of species of Orchidaceae.