The aim of this work was to investigate the bacteria-surface interactions occurring during the first hour of adhesion of marine Pseudoalteromonas NCIMB 2021 at the surface of 2304 lean duplex stainless steel in artificial seawater. A complete characterization of the biofilm and the passive film was performed coupling epifluorescence microscopy, scanning electron microscopy (SEM), x-ray photoelectron spectroscopy (XPS), and time of flight secondary ion mass spectrometry (ToF-SIMS). The coupling of XPS and ToF-SIMS analyses revealed that (1) protein and polysaccharide contents in the biofilm are similar in the presence or absence of nutrients, (2) the biofilm is mainly composed of proteins and the protein content is similar to the one of Tightly Bound EPS, (3) increased bacterial activity due to nutrients leads to chromium enrichment in the passive film in close vicinity to the bacteria.