Background: Hepatocellular carcinoma (HCC) grades second among cancer-related deaths. Rehmanniae Radix (RR) has been exposed to have numerous pharmacological properties. Objectives: To inspect RR leaves extracts pro-apoptotic activities on SMMC-7721 liver cancer cell lines. Materials and Methods: Anti-cancer effect of RR extracts in SMMC-7721 liver cancer cell lines was aged rate of cell apoptosis (AO/EB, 4′,6-diamidino-2-phenylindole [DAPI], and PI staining), ΔΨm (Rh-123 staining), intracellular Reactive oxygen species (ROS) (DCFH-DA staining), quantitative polymerase chain reaction (mRNA expressions of Bax, p53, Bcl-2, caspase-3,-9,-8, IL-6, PI3K/AKT, and mTOR). Results: It has been shown that RR leaf ethanolic extracts suggestively declined cell viability, augmented the rate of cell apoptosis (AO/EB, DAPI, and PI staining), and caused the potential for ΔΨm, and induced over-accumulation of intracellular ROS (DCFH-DA staining) in SMMC-7721 cells after 24 h of exposure. In SMMC-7721 cell lines, we have examined anti-cancer activity to scrutinize the underlying mechanism by assessing the mRNA expressions of Bax, p53, Bcl-2, caspase-3,-9,-8, IL-6, PI3K/AKT, and mTOR. Results displayed the significant inhibition of the apoptosis as stated above related proteins. Conclusion: RR extracts exhibited its potential anti-cancer activity against SMMC-7721 cells by prompting apoptosis-induced cell death and inhibiting PI3K/AKT/mTOR signaling.