Research studies on phytochemicals and antibiotics utilizing extracts from medicinal plants have proliferated in the past few years. The phytochemicals profiling, antioxidant capacity, antibacterial properties, antiproliferative potential and molecular docking studies of ethanolic extract of Onosma bracteatum was examined in this study. The extract prepared into ethanol (80%) and identify the presence of phytochemical compounds using GC-MS profiling. The antibacterial potential was investigated in relation to Staphylococcus aureus and Escherichia coli. Further to evaluate the anticancer potential of ethanolic extract of O. bracteatum, the antiproliferative assay (MTT cytotoxicity assay) was performed against Ln-18 (human malignant glioma cell line) and A-549 (Human lung cancer cell line) cells. Ethyl oleate, hexadecenoic acid and phytol were highest in GC-MS and docked against DNA gyrase B (E. coli), β-lactamase (S. aureus) and BCL-2 (apoptosis inhibitor). In antioxidant assay, 78% maximum inhibition of free radical scavenging activity was observed. The zone of inhibition was in the ranges from 9 mm (E. coli) to 20 mm (S. aureus). Furthermore, the ethanolic extract of O. bracteatum demonstrated antiproliferative activity against the cancer cell lines LN-18 and A-549 with growth inhibitions of 84% and 81%. The ethyl oleate was found to be best compound with -8.4 (kcal/mol) binding score in the molecular docking studies. The results showed that O. bracteatum is a good source of phytochemicals and antibacterial compounds with promising anticancerous potential.