Abstract: We investigated the medicinal vascular plant resources of Wuliangshan National Nature Reserve via field investigations, previous literature, and the collection and identification of plant specimens. Results showed there were 218 species of medicinal vascular plants belonging to 168 genera and 80 families, which accounted for 57.37% of total plant species. For growth forms, there were more species of herbs than of shrubs and trees. For medicinal vascular plant utilization, roots and whole herbs were the most commonly used part of the whole plant, accounting for 22.31% and 21.03% of the total, respectively. Analysis of medicinal vascular plant efficacy showed they were mostly used as antipyretics and antirheumatics and activating blood to resolve stasis. Assessment of the threatened status of the medicinal vascular plants showed that those of least concern accounted for 82.11% of the medicinal plants, which was much higher than that of other regions nation-wide and indicated that species in this region were well-preserved. Vascular plant species richness in the different functional zones of the reserve was:core zone > buffer zone > experimental zone, though the proportion of medicinal vascular plants was:core zone < buffer zone < experimental zone. Jaccard similarity analysis of medicinal vascular plants in different functional areas showed that the species in the core and experimental areas were extremely dissimilar, whereas the species in the core area and buffer area were slightly similar. This study shows that the establishment of protected areas plays an important role in the protection of the diversity of medicinal plants in Wuliangshan National Nature Reserve. Furthermore, we clarified, to a certain extent, the species composition and distribution of medicinal vascular plants, thus providing a reference for further development of protection and ethnobotany.