The present study was designed to investigated the chemical composition, aqueous extracts bioactive compounds and biological activities of Portulaca oleracea (PO) on human liver cancer (HepG2), human kidney cancer (Uo-31) cell lines and normal human lung cell lines (Wi-38) and compared with staurosparine (STA) as anticancer drug were evaluated. The obtained results the chemical composition of Portulaca oleracea powder was characterized with high protein, ash and total carbohydrate, which amounted in 15.07±0.01, 28.29±0.05 and 40.82±0.60% on D.W, respectively. The anticancer activity of extracts of Portulaca oleracea on three cancer cells line were exposed to various concentrations (0.4- 1.6- 6.3- 25- 100 μg /mL) for 24h. Percentages of cells viability were analyzed by (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)). From the obtained results showed that PO induced statistically significant (p < 0.001) decrease in the cell viability of HepG2 cells in a concentration dependent manner. The HepG2 cells exposed to PO at 100 μg/mL and lower concentrations were found to be cytotoxic. The cells viability of (STA/HepG2) was recorded as 32.70, 42.94, 51.40, 60.44 and 66.67% at concentrations 100, 25, 6.3, 1.6 and 0.4 μg/mL of PO, respectively. The cell viability (S1/HepG2) was recorded as 22.98, 29.56, 38.71, 46.89 and 59.29% at the same concentrations above, respectively. While, the decrease in the cell viability at 0.4, 1.6, 6.3, 25 and 100 μg/mL of PO extract was recorded to be 73.62, 65.27, 55.52,45.74 and 34.26% for (STA/UO-31) and 78.62, 70.51, 61.88, 52.49 and 38.24% for (S1/UO-31 cells), respectively. On the other hand, the normal cells line for (WI-38 cells) showed the heighest in the cell viability in a concentration dependent manner. The data from this study showed that exposure to extract of Portulaca oleracea resulted in significant cytotoxicity and inhibition of growth of the human liver cancer (HepG2) and human kidney cancer (UO-31) cells lines. From the obtained results, it can be concluded that Portulaca oleracea extracts were a significantly reduced cell viability of HepG2 and Uo-31 cells in a concentration-dependent manner. HepG2 cells were also found to be more sensitive to PO extract, according to the findings.