In this paper, the conformational changes of tryptophan (Trp) on the corroded 316 L stainless steel (SS) surface obtained under controlled simulated inflammatory conditions have been studied by Raman (RS) and Fourier-transform infrared (FT-IR) spectroscopy methods. The corrosion behavior and protective efficiency of the investigated samples were performed using the potentiodynamic polarization (PDP) technique in phosphate-buffered saline (PBS) solution acidified to pH 3.0 at 37 °C in the presence and absence of 10−2 M Trp, with different immersion times (2 h and 24 h). The amino acid is adsorbed onto the corroded SS surface mainly through the lone electron pair of the nitrogen atom of the indole ring, which adopts a more/less tilted orientation, and the protonated amine group. The visible differences in the intensity of the Fermi doublet upon adsorption of Trp onto the corroded SS surface, which is a sensitive marker of the local environment, suggested that a stronger hydrophobic environment is observed. This may result in an improvement of the corrosion resistance, after 2 h than 24 h of exposure time. The electrochemical results confirm this statement—the inhibition efficiency of Trp, acting as a mixed-type inhibitor, is made drastically higher after a short period of immersion.