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OBJECTIVE The objective of this study is to characterize breath-hold induced oxygenation changes in diabetic foot ulcers (DFUs) and develop an oxygenation flow index (OFI) to discern non-healing from healing DFUs. APPROACH The imaging approach utilizes an innovative breath-hold (BH) stimulus that induces vasoconstriction and measures for altering oxygenation flow in and around the tissues of DFUs and controls. The modified Beer-Lambert law was utilized to calculate hemoglobin-based spatio-temporal oxygenation maps in terms of oxygen saturation. RESULTS We found controls had synchronous BH induced oxygenation changes across the dorsal (OFI: 29.0%) and plantar (OFI: 57.6%) aspects of the foot. Non-healing DFUs, however, had less synchronous BH-induced oxygenation changes (OFI <28%). In addition, two complicated healing DFU cases, or cases with underlying issues or poor long term healing outcomes, were observed to have OFIs <28%. INNOVATION An OFI was developed to differentiate non-healing DFUs from healing DFUs using a single, non-contact, near infrared optical scanner for spatio-temporal oxygenation monitoring. The OFI has potential to provide immediate feedback on the microcirculation in DFUs, via hemoglobin-based oxygenation parameters. CONCLUSION A preliminary threshold (OFI<28%) could differentiate non-healing and complicated DFUs from healing DFUs. The overall oxygenation flow pattern was less synchronous (or the OFI value reduced) in the non-wound areas of the feet that were non-healing. In other words, the reduced OFI value (<28%) in the entire foot excluding the wound region is a possible indicator that the wound may not heal.