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In this study we discuss the cloning and expression of Entamoeba histolytica arginase (EhArg), an enzyme that catalyses the hydrolysis of l‐arginine to l‐ornithine and urea. l‐norvaline, a competitive inhibitor of E. histolytica l‐arginase, inhibits the growth of the parasite, which suggests that the catabolism of l‐arginine mediated by EhArg is essential. Nitric oxide (NO) is an antimicrobial agent that inhibits some key enzymes in the metabolism of Entamoeba histolytica. NO is synthesized by activated macrophages from l‐arginine, the substrate of NO synthase (NOS‐II). We show that E. histolytica inhibits NO mediated amoebicidal activity of activated macrophages by consuming l‐arginine present in the medium.