Iron plays a critical role in the production of activated oxygen species and the activity of chelated iron in the biological system depends on the chemical forms of the chelators. In the present study, we used ferric nitrolotriacetate (Fe-NTA, molar ratio of iron to chelators = 1:3), ferric ethylenediaminetetraacetate (Fe-EDTA, 1:3 complex) and ferric Desferal (Fe-Des, 1:1.1 complex) to see their "free" iron content in aqueous solutions in vitro and in the serum obtained after a single intraperitoneal injection of the chelates to rats (7.5 mg of iron/kg). "Free" iron was measured by the bleomycin-assay system. When Fe-NTA was dissolved in water, "free" iron increased linearly with total iron concentration up to 10 microM, whereas Fe-EDTA and Fe-Des showed no "free" iron with corresponding iron concentrations. When these three ferric chelates were dissolved in normal rat serum, "free" iron in Fe-NTA increased abruptly between 40 microM and 60 microM iron concentrations, then increased slowly up to 100 microM. Fe-Des did not show any "free" iron at comparable iron concentrations. Fe-EDTA had an intermediate "free" iron level in the serum. Among the ferric chelate complexes, Fe-NTA showed a much faster increase of and a higher content of "free" iron in the serum than the other two complexes after a single injection of the chelates into rats.(ABSTRACT TRUNCATED AT 250 WORDS)