Inflammation induced by oxidative stress causes further stress through positive feedback and may lead to various diseases. Antioxidant enzymes such as catalase can be useful therapeutic agents; however, their applicability is currently limited by their low stability. The aim of this study was to increase the stability of catalase by encapsulating this enzyme in nondegradable nanoparticles based on polyethylene glycol. Nanoparticle formation was confirmed using nanotracking analysis and transmission electron microscopy. The mesh structure of the nanoparticles allowed the passage of only very small molecules such as hydrogen peroxide (H2O2). The catalase encapsulated in the nanoparticles was stabilized because of isolation from destabilizing external factors such as proteases and was capable of decomposing H2O2 as determined by measuring the catalase activity. Upon introduction into HeLa cells, the nanoparticles protected the cells from oxidative stress caused by H2O2, which was confirmed by analyzing microscopy images and determining the number of live cells by cell staining. In the future, this method can ensure the continuous antioxidant activity of catalase in inflammatory cells and allow the retention of activities of other types of enzyme for extended periods within the cells, and also expected to be used as an intracellular sensor for monitoring enzyme function within cells.