Diabetic related ulcers are a common and severe complication of diabetes that can lead to severe health complications and prove fatal if mistreated or undiagnosed. Currently, there is lack of understanding regarding the underlying biochemical and physiological processes involved during chronic wound healing. The objective of this project was to understand and investigate the underlying factors in the wound environment that could be used to distinguish healing from non-healing diabetic foot ulcers. Bottom up proteomics was performed to analyse healing and non-healing representative pooled samples using SDS-PAGE and LC-MS/MS by AB SCIEX TripleTOF® 5600 System. The data analysis identified more than 400 proteins each from healing and non-healing samples, and 15 and 16 proteins unique to healing and non-healing respectively. A semi-quantitative approach involving spectral counting revealed more than 200 differentially regulated proteins in the samples. Proteins were grouped based on different functional properties and oxidative stress related proteins were selected for further analysis as there is growing evidence to suggest their vital role in the pathogenesis of diabetes related ulcers. The top 6 abundant proteins, catalase, peroxiredoxin 1, peroxiredoxin 2, superoxide dismutase-2 (SOD-2), S100-A9 and myeloperoxidase (MPO), were chosen on the basis of semi quantification results for further validation using enzyme-linked immunosorbent assay (ELISA).