The development of self-assembled nanocarriers for the encapsulation of hydrophobic antioxidants is of growing interest. Self-assembled amphiphilic chitosan conjugate nanocarriers that stabilize antioxidants were prepared based on the concept that both the nanocarrier and the antioxidant bear similar hydrophobic moieties able to establish hydrophobic interactions. This work describes the preparation and characterization of a system consisting of a palmitoyl chitosan conjugate and retinyl palmitate. Palmitic acid was coupled to chitosan using a carbodiimide-mediated coupling reaction, and two different palmitoyl chitosan conjugates were obtained by varying the coupling system. Palmitoyl chitosan conjugates self-assembled to form nanoparticles in aqueous medium varying in mean average diameter (Dh) between 200 and 437 nm. Retinyl palmitate–loaded nanoparticles were prepared by a solvent displacement method using dialysis, with loading efficiencies of 77.5% and 88.6%, loading contents of 12.6% and 14.6%, and Dh values of approximately 280 nm. The zeta potential (ζ) of all palmitoyl chitosan nanoparticle were above 25 mV, but ζ slightly increased in the retinyl palmitate–loaded nanoparticle. Antioxidant activity of loaded nanoparticles was confirmed using the 1,1-diphenyl-2-picryl-hydrazyl radical scavenging assay. The in vitro cytotoxicity of blank and loaded nanoparticles was determined using fibroblasts of human embryonic skin. All nanoparticles were not cytotoxic when they were tested with methylthiazol tetrazolium and lactate dehydrogenase tests. The obtained results suggest that the system has potential as a nanocarrier for dermal application. Additionally, the approach considered in this article can be expanded to other nanocarrier/antioxidant systems.