Magnesium hydroxide nanoparticles are widely used in medicinal and hygiene products because of their low toxicity, environment-friendliness, and low cost. Here, we studied the effects of three different sizes of magnesium hydroxide nanoparticles on antibacterial activity: NM80, NM300, and NM700. NM80 (D₅₀ = 75.2 nm) showed a higher bactericidal effect against Escherichia coli than larger nanoparticles (D₅₀ = 328 nm (NM300) or 726 nm (NM700)). Moreover, NM80 showed a high bactericidal effect against not only exponential cells but also persister cells, which are difficult to eliminate owing to their high tolerance to antibiotics. NM80 eliminated strains in which magnesium-transport genes were knocked out and exhibited a bactericidal effect similar to that observed in the wild-type strain. The bactericidal action involved physical cell damage, as confirmed using scanning electron microscopy, which showed that E. coli cells treated with NM80 were directly injured.