World-wide, Indonesia is ranked 17th in oral cancers, with deaths reaching 2.326 cases. Of the oral cancers, 90% are squamous cell carcinoma (HSC-3). Unfortunately, conventional cancer therapy still has many ill side effects. Therefore, pharmacologists have looked for natural ingredients to prevent the growth of oral cancer cells. One source is Cymbopogon citratus leaf. Research shows that the active compound of C. citratus leaf is a chemopreventive, doing so by increasing the production of re-active oxygen species (ROS) to induce apoptosis in cancer cells. The active compound of C. citratus leaf has low stability and solubility, so it is necessary to use an encapsulation matrix such as chitosan, and modify it into smaller particles to increase its effectiveness. Purpose is determining the effect of nano chitosan encapsulation of C. citratus leaf ethanol extract (NCECC) on the reactive oxygen species of HSC-3 tongue cancer cells. This study is divided into ten groups – without treatment, doxorubicin (positive control), hydrogen peroxide, nano chitosan and C. citratus leaf extract groups, and five groups of NCECC treatment – concentrations of 100%, 75%, 50%, 25%, and 12.5%, respectively. The 100% NCECC group showed the highest ROS concentration (p<0.05), compared to 75%, 50%, 25%, 12.5% NCECC groups, and the 100% NCECC was higher than the positive control group. NCECC is effective in inducing oxidative stress on HSC-3 through increased production of ROS. Moreover, the higher the encapsulation concentration given, the greater the increase in ROS production.