Ti-6Al-4V alloy has proven to be technically superior and cost-effective materials for a wide variety of aerospace, industrial, marine and commercial applications. The mechanical properties of Ti-6Al-4V are very sensitive to the microstructure obtained after the thermo-mechanical treatment. The duplex structures provide good tensile ductility, fatigue strength, resistance to microcrack growth and crack initiation, and are often used in demanding fatigue critical tasks. However, although Ti-6A-4V is considered to be reasonably resistance to chemical attack, severe problems can arise when it comes in contact with hydrogen-containing environments due to its susceptibility to hydrogen embrittlement. The objective of this paper is to investigate the absorption and desorption behavior of external hydrogen on a duplex-annealed Ti-6Al-4V alloy. While investigating the desorption profile, we seek to better understand the thermodynamics and the kinetic nature of the interaction between traps and hydrogen atoms, with specific emphasis on the investigation of the impact of these interactions on the microstructure of the studied aerospace applicative titanium alloy. In order to achieve these goals, thermal desorption spectroscopy (TDS) was applied and the data obtained from this analysis was supported by a variety of other experimental techniques, such as LECO hydrogen determinator, XRD and microstructure investigations by means of optic and electronic microscopy. Hydrogen was found to influence significantly the microstructure of the alloy. The process of hydrogen evolution was found to be a very complex process, being affected mainly by the phase transformations that may occur during the thermal analysis.