Effects of Certain Cations (Fe, Zn, Mg, and Ca) on Bacterial Endotoxins

The natural occurrence of cations Fe, Zn, Mg, and Ca in the lipopolysaccharide (LPS) of both the S and R forms of Shigella dysenteriae 1 was studied. LPS preparations were obtained either by phenol-water extraction (according to the method of Westphal et al., Z. Naturforsch. 7b:148-155, 1952) or by extraction of cells with hypertonic sodium chloride-sodium citrate (according to the method of Raynaud and Digeon, C. R. Acad. Sci. (Paris) 229:564-566, 1949), with subsequent chromatographic purification on Sephadex G200 and Sepharose 4B columns. The cation in highest concentration in the Westphal extract was Mg(2+) (as much as 30 mug/mg), and the lowest one was Fe (ca. 0.10 mug/mg). In LPS of the Raynaud type, the cation in highest concentration was Ca(2+) (as much as 13 mug/mg), and the lowest one was Fe (ca. 0.10 mug/mg). The effects of increasing and decreasing the concentrations of cations (Fe, Zn, Mg, Ca) upon the biological activity of the endotoxins was evaluated by using toxicity in mice and the Limulus test. It appeared that increased concentrations of Fe (chiefly of Fe(3+)) decreased the toxicity of the R form of LPS, whereas Mg(2+) decreased the toxicity of the S form. After prolonged dialysis of LPS preparations against deionized water, there was no consistent relationship between toxicity as determined in white mice and with the Limulus test.