Controlled drug release system is one of the most favorable technique of novel drug delivery system owing to its reproducibility and ease of formulation. Nanotechnology is very useful for controlling the drug release and thus improving the pharmacokinetic and pharmacodynamic properties of the drug. The technique improves patient compliance by reducing both dose and the frequency of administration and thus minimizing the local as well as systemic toxic effects. The aim of the present research work was to formulate and evaluate nanoparticles of Rutin by using the Emulsion solvent evaporation method. Sustained release nanoparticles of Rutin were prepared to increase the drug residence time in gastrointestinal tract and thus improving the bioavailability of drug. The nanoparticles were prepared by using Chitosan and Carbopol 940 as polymers. Different formulations were prepared with varying concentrations of Chitosan and Carbopol 940 in order to achieve the optimum particle size and maximum encapsulation efficiency. The particle size of nanoparticles was found to be in the range of 0.181±0.051 nm to 0.390±0.101 nm. Drug encapsulation efficiency ranged between 58.1±0.651 percent to 82.9±1.216% with controlled drug release up to 99.29% in phosphate buffer pH 6.8, 12 hrs. FT-IR studies showed that the drug and polymers were compatible. The results of Nanoparticles indicated that optimized formulation exhibited excellent properties. Nanoparticles in the doses of 50 mg/kg, 100 mg/kg, and 200 mg/kg were used in wistar rats of either sex. The oxidative stress was produced by overdose of acetaminophen and estimation of serum concentration of various enzymes such as malonaldehyde (MDA), superoxide dismutase (SOD), reduced glutathione (GSH), and catalase (CAT) were measured by standard biochemical methods. Silymarin (100 mg/kg) was used as a standard drug for assessment of antioxidant status. When compared with the standard antioxidant silymarin, Nanoparticles did not exhibit antioxidant activity in terms of MDA level reduction. However, it significantly increased serum levels of the antioxidant enzymes (SOD, GSH, and CAT) exerting a potent antioxidant effect in a graded manner. The observed results suggest that Rutin loaded Nanoparticles could be a potential source of antioxidants. However, further studies are required to explore this therapeutic property of Rutin. Keywords: - Nanoparticles, Chitosan, Carbopol 940, Rutin and Emulsion solvent evaporation method and antioxidant.