Superoxide anion generation in effective and ineffective soybean root nodules

Superoxide anion generation was measured in soybean root nodules through the autoxidation of epinephrine, resulting in the formation of adrenochrome. Superoxide anion production was initially high in effective nodules but it rapidly declined following an increase in nitrogen fixation capacity. Superoxide anion levels sharply increased after a drop in the nitrogen fixation rate and reached their maximum rates between 35 and 42 days post-inoculation. Superoxide anion rate of formation was 2 to 4 times higher in the nodules than their respective adjacent root tissue. The rate of epinephrine autoxidation was partially dependent on EDTA and NAD(P)H. Superoxide anion production was inhibited between 80 to 90 percnt; by 100 mmol/L of HEPES, MES, MOPS, Tiron, thiourea and two units of superoxide dismutase whereas BSA, DMSO, mannitol, methionine, phenylalanine and sodium benzoate (quenchers of hydroxyl radicals) caused only a partial suppression of superoxide anion formation. On the other hand, both sodium azide and mercapto-ethanol, stimulated O2*− production.