Echinops spinosissimus Turra subsp. bovei (Asteraceae) is a medicinal plant in western Algeria. Traditionally, roots and inflorescences are employed as hypertensive agents and in the treatment of hemorrhoids. The current study evaluates the chemical composition, antioxidant, antimicrobial, and wound-healing properties of the root methanolic extract from E. spinosissimus subsp. bovei. The content of total phenolics, flavonoids, and tannins was determined. In addition, the phenolic profile was typified. The studied plant extract resulted in being primarily composed of Apigenin, Kaempferol, and their derivatives. The total phenolic content was equal to 95.31 ± 2.90 mg GAE/g DW, while the number of flavonoids and condensed tannins was 16.01 ± 0.16 mg CE/g DW and 8.30 ± 0.65 mg CE/g DW, respectively. The methanolic extract was found to exhibit antioxidant activity towards the DPPH radical, with an IC50 of 7.99 ± 0.28 mg/mL and a TAC of 30.30 ± 0.54 mg AAE/g DW, as well as an antibacterial effect, especially against P. aeruginosa. No significant wound-healing property was observed, even though the histopathological observations showed enhanced wound-healing quality. According to our evidence, E. spinosissimus could represent a source of phytochemicals with potential beneficial effects for human health in terms of antioxidant and antibiotic properties, although further investigations on this species are needed.