Metabolic dysregulation is a hallmark of hepatocellular carcinoma (HCC). AMPK is a crucial hub of metabolic regulation during cancer progression. We show that phytochemical Levo-tetrahydropalmatine (THP) activates AMPK-dependent autophagy to downregulate the mitochondrial respiration and glycolysis. Consequently, THP significantly decreased cell viability in two HCC cell lines, BEL-7402 and SMMC-7721. Similarly, NOX4 inhibitor diphenyleneiodonium chloride (DPI) induces concomitant downregulation of the mitochondrial and glycolytic metabolism. In contrast to THP, cells are less sensitive to proliferation inhibition induced by DPI treatment as compared to THP treatment did. Combined treatment of THP and DPI was found to be more efficacious in killing cancer cells than either of the agents treated individually. Indeed, the co-operative effect by the THP-DPI combination improves the pro-apoptotic activity in response to the energy depletion as outlined by a drastic decrease in ATP levels. Therapeutic regime significantly reduced the tumor growth in mice. Importantly, this is realized without causing systemic toxicity to other organs. Collectively, our work shows that the combinatorial therapy of autophagy activator THP and NOX4 inhibitor DPI may be considered as a therapeutic avenue against HCC. © 2022 Elsevier Ltd