Intracellular free iron of Escherichia coli was determined by whole-cell electron paramagnetic resonance spectrometry. Ultrahigh pressure (UHP) increased both intracellular free iron and cell lethality in a pressure-dose-dependent manner. The iron chelator 2,2'-dipyridyl protected cells against UHP treatments. A mutation that produced iron overload conditions sensitized E. coli to UHP treatment.