Updated project metadata. The clinical assessment of wound infection and related complications is challenging and the development of objective methods to measure wound status and predict healing outcomes is therefore essential. As bacterial protease activities play important roles in infection, analysis of changes in the wound peptidome by individual bacterial enzymes could provide insight into proteolytic events occurring in infected wounds, and may aid in the discovery of biomarkers. Wound infection was mimicked by incubating sterile acute wound fluids ex vivo with the bacterial proteases Pseudomonas aeruginosa LasB and Staphyloccocus aureus V8. Using a peptidomics approach, we characterized the low-molecular-weight peptidome of the digested samples and identified over 100 protein targets for each enzyme. Additionally, we found enzyme-specific peptides and digestion patterns.