Stomach contents of mice fed on a standard rodent breeding diet contained 29-733 microM-soluble nonhaem-iron. A very variable percentage (3-100, mean 49.3 (SE 4.7), n 37) of this Fe was rapidly (half-life less than 1-2 s) available for chelation by the strong Fe(II) chelator ferrozine, with little or no further Fe being available on addition of ascorbate. Ferrozine-available Fe could be detected in the duodenal lumen at concentrations up to 60 microM in vivo and after in vitro neutralization of stomach contents. No significant changes in quantity of stomach ferrozine-available Fe or soluble non-haem-Fe occurred in mice with adaptive enhancement of Fe absorption induced by chronic hypoxia. Electron paramagnetic resonance (e.p.r.) spectroscopy of the soluble portion of mouse stomach contents demonstrated a g = 4.3 signal (rhombic Fe(III)) equivalent to up to 20% of soluble non-haem-Fe. The signal was unaffected by addition of excess ferrozine and increased on subsequent neutralization, suggesting redistribution of Fe from other e.p.r.-silent species. Solutions of Fe-nitrilotriacetate (NTA) (a synthetic Fe chelate used as a bioavailable, model Fe solution) were found to contain both rapidly and slowly ferrozine-available Fe (after addition of ascorbate) depending on pH, NTA:Fe ratio and the presence of Ca(II) ions. Fe-ascorbate mixtures (a model solution for Fe absorption studies) also contained ferrozine-available Fe. These results suggest the presence of Fe(II), rhombic Fe(III) and other e.p.r.-silent Fe species in the soluble fraction of mouse stomach contents. The ferrozine-available (Fe(II)) fraction is not limited by the reducing power in the diet, but by binding to ligands. Neutralization with bicarbonate leads to a loss of ferrozine-available Fe and increase in rhombic Fe(III) at the expense of both ferrozine-available and other e.p.r.-silent Fe species. The ferrozine-available Fe in mouse stomach and duodenal lumen can be related to Fe species present in model solutions used for in vitro studies of mucosal uptake mechanisms.