The ethnobotanical survey of Euclea divinorum Hiern (Ebenaceae) was conducted on Soqotra Island, Yemen. The root bark is used to treat mouth, dental, dermal and blood diseases in the traditional medicine of the island. The study is the first report about the effect of the plant root barks against six human pathogenic fungi. The non-polar dichloromethane extract of Euclea divinorum root bark showed stronger antifungal activities compared to polar direct and sequential methanolic extracts. These extracts showed significant broad antifungal activity against Absidia corymbifera, Aspergillus fumigatus, Candida krusei, Microsporum gypseum, Mucor sp. and Trichophyton mentagrophytes compared to the standard antibiotic drug nystatin. Thin-layer chromatography (TLC) revealed the presence of the naphthoquinones in the extracts. The results showed an extraction process to separate most antifungal naphthoquinones from the root bark by using non-polar solvent dichloromethane, while flavonoids remained in the polar methanolic extracts; therefore, the polar sequential and direct methanolic extracts recorded higher antioxidant activity than the non-polar extracts with less toxicity. The biological and chemical results identified the presence of antifungal and antioxidant constituents in the root bark and confirm its traditional use in Soqotra Island as crude powder to treat dental and dermal diseases and to clean teeth. Moreover, our results are compared with literature review on ethnobotany and phytopharmacology of E. divinorum to present a medicinal monograph about the plant.