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The aim of the present study was to identify the active principle from Catharanthus roseus leaf using larvicidal bioassay against three mosquito species viz. Aedes aegypti, Culex quinquefasciatus, and Anopheles stephensi. Preliminary studies of the three successive extracts such as hexane, chloroform, and methanol against Ae. aegypti larvae showed that the chloroform extract was more active with LC 50 and LC 90 values of 40.09 ppm and 189.15 ppm respectively. Bioassay guided fractionation of the active chloroform extract resulted in the isolation of a triterpenoid (ursolic acid) as the active constituent. Three derivatives acetate, formate, and benzoate were prepared using this, and they were tested for their larvicidal activity against three mosquito species. The acetyl derivative was highly active against all the three species compared to the parent compound ursolic acid; the activities of benzoate and formate were higher than ursolic acid when tested against Cx. quinquefasciatus. This is the first report related to ursolic acid from C. roseus with mosquito larvicidal activity. The pure compound could be considered for medicinal and other pharmacological applications in future., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)