Since ancient times, plants have been used as a valuable source for treating serious diseases. Three different plants, namely Aubrieta pinardii , Johrenia dichotoma , and Malabaila dasyantha were used and extracted using the extraction solvents ethyl acetate (EA), methanol (MeOH), and water. Antioxidant activity was determined using several in vitro assays. Subsequently, the enzyme inhibition activities of the extracts were evaluated against cholinesterases (AChE and BChE), tyrosinase, amylase, and glucosidase. The antibacterial and antifungal effects of the extracts were assessed. Methanol extracts of A . pinardii , J . dichotoma , and M . dasyantha yielded the highest total flavonoids (TFC) compared to EA and water extracts. The analysis using HPLC-ESI-Q-TOF-MS identified various classes of phytochemicals within the species examined. Aubrieta pinardii extracts were abundant in glucosinolates, whereas J. dichotoma and M. dasyantha extracts were characterized by the presence of coumarins and furanocoumarins. The EA extracts exhibited greater inhibition of microorganism growth compared with MeOH and water extracts. Notably, J. dichotoma showed similar or superior antifungal activity against A. niger (MIC/MFC 0.25/0.50 mg/mL) and M. dasyantha demonstrated comparable efficacy against T. viride (MIC/MFC 0.50/1.00 mg/mL) and P. verrucosum var. cyclopium (MIC/MFC 0.125/0.25 mg/mL) when benchmarked against the reference drug ketoconazole. These results provide valuable information about A . pinardii , J . dichotoma , and M . dasyantha chemical profiles and underline the importance of selecting appropriate solvents for the extraction of metabolites from plant matrix for potential applications in the treatment of various diseases.