Medicinal plants used in Ethiopian traditional practices are an important source of bioactive compounds with potential antiparasitic properties. Uvaria scheffleri (Annonaceae), Zanthoxylum chalybeum, and Vepris dainelli (Rutaceae) have long been employed to manage infectious conditions; however, their essential oils have not been previously examined for activity against Anisakis larvae. Essential oils were extracted from the roots of U. scheffleri and the fruits of Z. chalybeum and V. dainelli by hydrodistillation. Chemical profiling was performed using gas chromatography coupled with mass spectrometry. Larvicidal efficacy against third-stage (L3) Anisakis larvae was evaluated in vitro using marinated systems, while cytotoxic effects were determined on VERO cell lines via the MTT colorimetric assay. The extraction process yielded 0.5% essential oil from U. scheffleri, whereas higher yields were obtained from Z. chalybeum (2.7%) and V. dainelli (2.0%). GC–MS analysis identified 58, 18, and 20 compounds in the respective oils, accounting for over 97% of their total compositions. Tricyclo[5.3.0.0(3,9)]decane was the predominant constituent in the oils of Z. chalybeum and V. dainelli, comprising 82.8% and 69.8%, respectively, and is reported here for the first time. All tested oils induced rapid, dose-related mortality in Anisakis L3 larvae, with complete lethality observed within 3 h for Z. chalybeum and within 5 h for U. scheffleri and V. dainelli at a concentration of 5%. Cytotoxicity assays revealed moderate effects on VERO cells, with IC₅₀ values of 65.46, 83.88, and 96.82 µg/mL, respectively. The findings demonstrate that the essential oils of these Ethiopian medicinal plants possess strong larvicidal activity against Anisakis L3 larvae while exhibiting relatively low cytotoxicity. These properties support their potential development as safe, plant-derived larvicidal agents for the prevention and control of human anisakidosis.
