We postulated omeprazole inhibition of the neutrophil proton pump, impairing phagocytosis and phagolysosomal acidification. Neutrophils from healthy human beings were treated with omeprazole prodrug 0.5 mM/l or acid activated omeprazole 0.5 mM/l, then incubated with killed Saccharomyces cerevisiae stained with bromcresol purple. Wet mounts were done at 10, 30 and 60 minutes. Percent neutrophils phagocytosing, percent yeast phagocytosed, and yeast per phagocytosing neutrophil were significantly decreased in acid activated omeprazole compared to controls and omeprazole prodrug. In contrast, percent acidification of intracellular yeast was significantly lower in both omeprazole prodrug and acid activated omeprazole compared to controls. Over time, control neutrophils showed an increase in percent yeast phagocytosed and yeast per phagocytosing neutrophil. When treated with acid activated omeprazole, the percent of neutrophils phagocytosing progressively decreased over time. We observed 1) omeprazole prodrug does not inhibit neutrophil phagocytosis but does inhibit phagolysosomal acidification, whereas 2) acid activated omeprazole inhibits both neutrophil phagocytosis and phagolysosome acidification. We conclude that omeprazole impairs these neutrophil functions in vitro.