The superoxide dismutase mimic Mn(II/III) tetrakis (1-methyl-4-pyridyl) porphyrin (Mn(II/III)TMPyP) was examined for its superoxide radical (O2.-)-scavenging ability in cultured mammalian cells. Mn(III)TMPyP (< 5 microM) added to culture media relieved growth inhibition and decreased the inactivation of the O2(.-)-sensitive enzyme aconitase in cells exposed to the O2(.-)-generating phenazine pyocyanine. Treatment of cells with Mn(III)TMPyP did not measurably affect cellular O2.- production as revealed by rates of cyanide-resistant respiration with or without added pyocyanine. In contrast, Mn(II/III)TMPyP enhanced O2.- production in cells when the redox-active naphthoquinone menadione was present as measured by both increased cyanide-resistant respiration rates and aconitase inactivation. In vitro, Mn(II/III)TMPyP catalyzed the oxidation of ascorbate, and menadione enhanced this effect. Mn(III)TMPyP did not protect aconitase when O2.- production was elicited in mitochondria by antimycin A and the uncoupler carbonyl cyanide p-trifluoromethoxyphenylhydrazone. The results support a reductant-O2.-:oxidoreductase mechanism for O2.- scavenging by Mn(II/III)TMPyP in the mammalian cytosol as proposed for its action in Escherichia coli, but also indicate that Mn(II/III)TMPyP can either scavenge or produce O2.- in cells depending upon the prevailing pathways of Mn(II/III)TMPyP oxidation-reduction.