The relation between Olea europaea subspecies cuspidata and subspecies europaea in Saudi Arabia were investigated for determination of the bioactive compounds in olive leaves. The antioxidant and total phenolic content were demonstrated by HPLC/MS. Also, the antimicrobial agar gel well diffusion activity was done for ethanolic extracts. The study identified a great number of phenolic compounds out of which some anticancer compounds were identified. The content of olive leaves extract differs according to habitat (either wild or cultivated). The ethanolic extract of both types has high phenolic content (21.3 to 22.6 mg GA/gdw) and antioxidant activity (71% to 57%) for wild and cultivated leaf extracts, respectively. The phenolic profiles revealed the presence of plenty of flavonoids and phenolic compounds. The major polyphenol in the extracts is naringenin, which appears at peaks 8 with concentrations of 21.93 and 17.35 µg/mL for cultivated and wild leave respectively, which retention times 14.99 and 14.98. The antimicrobial activity showed that the leaves extract have strong antimicrobial activity against Bacillus subtilis, with the highest inhibition zones (28 and 26 mm) for wild and cultivated leaf extracts, respectively. This is considered as valuable data about the potential for industrial mass production of polyphenol.