Bactericidal properties of hydrogen peroxide and copper or iron-containing complex ions in relation to leukocyte function

Various combinations of hydrogen peroxide, reductant (ascorbic acid and superoxide ion), and copper or iron salts and their coordination complexes were examined to determine their cytotoxicity toward several bacteria with diverse metabolic capabilities and cell envelope structures. Four sets of bactericidal conditions were identified, comprising: (1) high concentration levels (5–100 mM) of H2O2 in the absence of exogenous metal ions and reductant; (2) ferrous or ferric coordination complexes plus enzymatically generated O2·− and H2O2 at relatively low steady-state concentration levels; (3) cupric ion plus low concentration levels of H202 (1 μM-1 μM) and ascorbate (10μM-4 mM); (4) cuprous ion (or cupric ion plus ascorbate) in the absence of 02 and H202. Rates of losses in viabilities increased proportionately with increases in the concentration of H2O2 in metal-free environments and with each of the components in the Cu2+/ascorbate/H2O2 bactericidal assay system. Oxidant levels required for equivalent killing increased with increasing cell densities of the bacterial suspensions over the range investigated (2 × 107-2 × 109 cfu/ml). Other experimental conditions or other combinations of reagents, most notably Fe3+/ascorbate/H2O2 systems, did not generate bactericidal environments. The patterns of response of the three organisms tested, Streptococcus lactis, Escherichia coli, and Pseudomonas aeruginosa, were similar, suggesting common bactericidal mechanisms. However, preliminary evidence suggests that the lethal lesions caused by the various bactericidal conditions are distinct. As discussed, each of the four bactericidal conditions could conceivably be attained within the phagosomes of leukocytes, although none has as yet been identified.