Electron paramagnetic resonance (EPR) spectroscopy represents an established tool to study properties of microenvironments, e.g. to investigate the structure and dynamics of biological and artificial membranes. In this study, the partitioning of the spin probe 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) in ex vivo human abdominal and breast skin, ex vivo porcine abdominal and ear skin as well as normal and inflammatory in vitro skin equivalents was investigated by EPR spectroscopy. Furthermore, the stratum corneum (SC) lipid composition (as determined by high-performance thin-layer chromatography), SC lipid chain order (probed by infrared spectroscopy) and the SC thickness (investigated by histology) were determined in the skin models. X-band EPR measurements have shown that TEMPO partitions in the lipophilic and hydrophilic microenvironment in varying ratios in different ex vivo and in vitro skin models. Ex vivo human abdominal skin exhibited the highest amount of TEMPO in the lipophilic microenvironment. In contrast, the lowest amount of TEMPO in the lipophilic microenvironment was determined in ex vivo human breast skin and the inflammatory in vitro skin equivalents. Individual EPR spectra of epidermis including SC and dermis indicated that the lipophilic microenvironment of TEMPO mainly corresponds to the most lipophilic part of the epidermis, the SC. The amount of TEMPO in the lipophilic microenvironment was independent of the SC lipid composition and the SC lipid chain order but correlated with the SC thickness. In conclusion, EPR spectroscopy could be a novel technique to determine differences in the SC thickness, thus suitably complementing existing methods.

Charité Universitätsmedizin Berlin Corporate Member of Freie Universität Berlin Humboldt Universität zu Berlin and Berlin Institute of Health Department of Dermatology Venerology and Allergology Center of Experimental and Applied Cutaneous Physiology Charitéplatz 1 10117 Berlin Germany

Charité Universitätsmedizin Berlin Corporate Member of Freie Universität Berlin Humboldt Universität zu Berlin and Berlin Institute of Health Department of Dermatology Venerology and Allergology Center of Experimental and Applied Cutaneous Physiology Charitéplatz 1 10117 Berlin Germany; Institute of Pharmacy Freie Universität Berlin Königin Luise Straße 2 4 14195 Berlin Germany

Department of Physics Institute of Experimental Physics Freie Universität Berlin Arnimallee 14 14195 Berlin Germany

Institute of Pharmacy Freie Universität Berlin Königin Luise Straße 2 4 14195 Berlin Germany; University of British Columbia Faculty of Pharmaceutical Sciences 2405 Wesbrook Mall V6T1Z3 Vancouver Canada

Skin Barrier Research Group Faculty of Pharmacy Charles University Akademika Heyrovského 1203 500 05 Hradec Králové Czech Republic

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