The current research aims to identify the chemical constitutes of the essential oils of the extracted Carduus pycnocephalus by Gas Chromatography-Mass Spectroscopic analysis (GC-MS) and to assess the biological profiles of the plant and its botanical ingredients as a precise antioxidant, and anticancer, as well as, antimicrobial agents. The extraction process of the medicinal plant by methanol provided a possibility to extract and identify the polar chemical constitutes that have the most effective categories of components. The DPPH antioxidant potency of the flower extract is the most potent with IC50 = 30.69 mg L-1 followed by leaves (IC50= 32.78 mg L-1), stem (IC50= 41.31 mg L-1), and root (IC50= 46.84 mg L-1). Leaf, and flower extracts revealed the most potent activities than the antibiotic standards against E. coli, S. typhimurium, and B. cereus species with inhibition zones ranged from 20-26 mm. Also, the extracted C. pycnocephalus revealed a moderate cytotoxic effect against hepatocellular carcinoma (HepG2) tumor cell line using MTT assay with IC50= 46.2 µg mL-1. The experimental interpretations inveterate the potential of C. pycnocephalus extract indicated its biological impacts as antioxidant, antibacterial, and moderate cytotoxic agents that provided the ease of using it in cancer therapy.