The sodium consumption in many countries is too high, which results in increased risk for hypertension, cardiovascular diseases, stroke and premature death. Inhomogeneous sodium distribution using layering is a viable way to reduce sodium in bread that normally contains a lot of sodium. Prevention of sodium migration during production and storage is important for the function of this approach. Furthermore, the distribution of sodium between starch and gluten influences their properties. The spatial distribution of sodium was investigated at high resolution using combinations of X-ray fluorescence microscopy (XFM), scanning transmission X-ray microscopy (STXM), light microscopy (LM), scanning electron microscopy with energy-dispersive X-ray spectroscopy (SEM-EDX) and image analysis. Reference breads and layered bread samples were baked with one salt-free layer and one layer containing 3.6 wt% sodium chloride salt. The obtained results showed that the concentration of sodium is higher in the starch phase than in the glutenphase and that sodiummigrates across the layer interface from the salt-containing to the salt-free layer. The ratios betweenthe sodium concentration in the starch and gluten phases were dependent on the sodium concentration across the interfaces. Furthermore, magnesium and phosphor signals in bread yeast cells were observed using XFM.