Lansoprazole is a gastric parietal cell proton pump inhibitor that is also active against Helicobacter pylori in vitro. We aimed to investigate further the mechanism of its antimicrobial effect. The antimicrobial activity of lansoprazole and of its sulfenamide, a rearrangement product occurring spontaneously in acid environments, was studied by determining the MICs and MBCs for 11 cytotoxic and eight non-cytotoxic H. pylori strains and by measuring the rapidity of bacterial killing. The MIC90 and MBC90 were 2.5 mg/L and 10 mg/L, respectively, both for lansoprazole and for its sulfenamide. Cytotoxic strains were as susceptible as non-cytotoxic strains. The sulfenamide exhibited faster bactericidal activity. Lansoprazole did not inhibit the toxin-induced vacuolization of HeLa cells by a cytotoxic strain, hence its anti-H. pylori activity does not depend on inhibition of a v-ATPase-mediated, toxin-induced activity. Sulfenamide formation is likely to occur in vivo in the gastric environment, thus enhancing the bactericidal activity of the drug. Lansoprazole is likely to be useful, in association with antibiotics, in the treatment of H. pylori infection regardless of the cytotoxicity of the infecting strain.