Chlorogenic acid (CGA) is an important natural polyphenolic antioxidant, composed of an ester of caffeic and quinic acid. The present work investigates the effect of cholesterol (Chol) on the action of CGA on membrane properties and its relationship with antioxidant capacity in systems propense to lipid peroxidation. The evaluation of CGA's interactions with membrane components and its influence on mechanical and electrical properties was conducted in unsaturated phospholipid monolayers, both in the presence and absence of Chol. This analysis included measurements of changes in surface pressure and dipole potential to assess this interaction. Furthermore, the apparent affinity of CGA in lipid vesicles, with and without Chol, was determined through zeta potential (ζ) measurements. These studies demonstrate that CGA exhibits an antagonistic effect with Chol on the membrane's biophysical properties due to specific interactions, directly influencing adsorption and antioxidant capacity. Consequently, CGA delays lipid peroxidation, preserves the integrity of lipid vesicles, and offers enhanced protection in the presence of Chol. These results contribute to improving our understanding of membrane protection against lipid peroxidation and highlight CGA's potential as an effective antioxidant in lipid systems.