Recently, there has been a growing awareness in scientific circles regarding the use of ethnomedicines for treatment cancers. This attention has largely arisen due to the limited selectivity of current chemotherapy and radiotherapy usages. In this study, aerial parts of Rosmarinus Officinalis from three distinct geographic locations were extracted using ethyl acetate. The crude extracts underwent evaluation for polyphenolic content and antiproliferative activity using techniques such as ELISA and xCELLigence assay. Total phenolics and flavonoids were quantified, with total phenolic yields ranging from 73.75 to 167.91 μg EAG/mg, and flavonoid yields varying from 66.2 to 93.1 μg EQ / mg. Furthermore, the extracts were tested against two cancer cell lines, C6 (rat brain tumor) and Hela, revealing moderate activity, particularly at a concentration of 250 μg/mL. These findings underscore the impact of climatic factors on the production of secondary metabolites, thus influencing biological activity.