The medical application of nanomaterials is growing fast. Amongst the most widely used, silver nanoparticles are antimicrobial agents whose key application is the care of burns and chronic wounds. Still, their absorption, distribution, metabolism and excretion behaviour in vivo has not yet been systematically investigated. We collected full-profile specimens of skin from four hospital patients with mid-to-deep thickness burns or equivalent skin wounds, treated with dressings containing silver nanoparticles or silver sulfadiazine. Synchrotron radiation μXRF/μXANES and laser ablation-ICP-MS were used to provide the first semi-quantitative/high resolution direct information on the spatiotemporal distribution and speciation of silver in vivo. The metal was rapidly released onto the wound surface, followed by a significant structure-dependent penetration into the damaged tissues. This was accompanied by sequential processes of metallic silver dissolution, chloride complexation, change to metal-thiol protein complexes, and final mobilization into deeper skin layers towards the vascular networks. Complete local clearance of silver was observed after 12 days of treatment in the case of full healing. The results provide a complete insight into the dynamics of silver in real human wounds, and a new basis for the design of innovative silver nanomaterials with optimal antibacterial efficacy and minimized risk for the patient.