Infection with Epstein-Barr virus (EBV) and its encoded latent membrane protein 1 (LMP1) play oncogenic roles in Burkitt's lymphoma (BL). Flow cytometry was used to measure cellular reactive oxygen species (ROS) concentrations, and cellular lactate generation and diphenylene iodonium (DPI) cytotoxicity were determined by analyzing lactate concentrations and cell viability. We also measured NAD(P)H oxidase (NOX) activity. Reverse transcriptase PCR and qPCR assays were used to analyze LMP1 levels, and protein expression was measured by immunoblotting. In the present study, EBV was able to induce NOX activity and ROS generation in the BL cells. Inhibition of NOX activity by DPI suppressed ROS levels and elevated lactate levels. DPI treatment first resulted in a G2-M phase cell cycle arrest and then induced significant apoptosis. Immunoblot analysis demonstrated that DPI suppressed the expression of c-Myc and Cdc25A within 6 h, which may have caused the cell cycle arrest. Collectively, these findings indicate a close relationship between EBV infection and NOX activation, permitting a deeper understanding of ROS inhibition in cell cycle regulation and providing a novel therapeutic target for BL treatment.