The effect of hexametaphosphate (HP), a long-chain polyphosphate known to have an antibacterial (bacteriostatic) action on Staphylococcus aureus through its action on cytoplasmic membranes, was studied. Using spheroplasts prepared from this bacterium, HP was observed to stimulate the leakage of nucleic acid-related substances, nucleic acids and proteins from the spherotoplasts in 1.2mol/L sucrose solution. In addition, using cytoplasmic membranes isolated from the cells, it was found that HP slowly released proteins in small amounts and rapidly released magnesium in large amounts from the membranes. The results indicated that HP acts on the cytoplasmic membranes. These findings suggest that the influence of HP on the cytoplasmic membranes of S. aureus plays an important role in its action on this bacterium.