Salvia virgata as an ethnomedicinal plant comprehends a variety of efficient active ingredients and shows diverse pharmacological actions, such as anti-proliferative, anti-inflammatory, and antimicrobial effects. In this study we aimed to determine the biological content of the ethanolic extract of S. virgata and to determine its possible pharmacological effects. The broth microdilution technique was carried out for determining antimicrobial activities, and the test microorganisms included E. coli, S. aureus, B. cereus, P. aeruginosa, C. albicans, and C. tropicalis. DPPH and ABTS methods were used to detect antioxidant activity. The XTT cell viability test was utilized to assess the antiproliferative activity of the ethanolic extract of S. virgata on L929 and MDA-MB-231 cell lines. S. virgata exhibited reasonable antimicrobial effects against E. coli (0.312 mg/mL) and S. aureus (0.312 mg/mL). DPPH and ABTS IC50 values were determined 291.58 ± 0.004 μg/mL, 16.74 ± 0.007 μg/mL respectively. S. virgata ethanolic extract TPC and TFC were observed 283.35 ± 10.4 mg GAE/g and 13.37 ± 1.6 mg QE/g, respectively. The extract was screened against α-amylase and α-glucosidase, AChE, and BChE enzymes, and inhibition activity was determined 75.73%, 62.72%, 67.19%, 3.18% respectively. The extracts did not significantly affect the L929 cell viability, while MDA-MB-231 remarkably reduced cell viability (IC50 = 0.118 mg/mL). The ethanolic extract of S. virgata can be considered as a potential therapeutic agent in the treatment of different pathological conditions due to its antimicrobial, antioxidant, and anticancer effects.