OBJECTIVE Graphene oxide-based nanotechnology has aroused a great interest due to its applications in the biomedical and optoelectronic fields. The wide use of these materials makes it necessary to study its potential toxicity associated with the inhalation of Graphene Oxide (GO) nanoparticles and its interaction with the lung surfactant. Langmuir monolayers have proven to be an excellent tool for studying the properties of the lung surfactant and the effect of intercalation of nanoparticles on its structure and properties. Therefore, to know the origin of the phospholipids/GO interaction and the structure of the lipid layer with GO, in this work we study the effect of the insertion of GO sheets on a Langmuir film of 1,2-Dipalmitoyl-sn-glycerol-3-phosphocholine (DPPC). METHODS Surface pressure-area isotherms, Neutron (NR) and X-ray Reflectivity (XRR) and Grazing Incidence X-ray Diffraction (GIXD) measurements of hydrogenated and deuterated DPPC monolayers with and without GO have been carried out. RESULTS The results outline a strong interaction between the GO and the zwitterionic form of DPPC and prove that GO is in three regions of the DPPC monolayer, the aliphatic chains of DPPC, the head groups and water in the subphase. Comparison between results obtained with hydrogenated and deuterated DPPC allows concluding that both, electrostatic attractions, and dispersion forces are responsible of the interaction GO/DPPC. Results also demonstrated that the insertion of GO into the DPPC aliphatic chains does not induce significant changes on unit cell of DPPC.