Two species of Leishmania (L), L. tropica and L. major, are among the main causative agents of cutaneous leishmaniasis. Arginase (ARG) is an essential enzyme for cell growth, thus an attractive drug target. In this study, we tried to survey the inhibitory impact of ARG by nor‐NOHA (N‐ω‐hydroxy‐L‐nor‐arginine) on in vivo infection caused by L. tropica. BALB/c mice were inoculated with L. tropicaEGFP‐LUC (Ltrop) or L. majorEGFP‐LUC (Lmj) and then were treated by nor‐NOHA. ARG inhibitor only indicated a delay in generation of a cutaneous lesion in inoculated footpad with nor‐NOHA‐Ltrop and nor‐NOHA‐Lmj. ARG activity has been significantly reduced in nor‐NOHA‐Ltrop group. In this group, ARG activity inhibition correlated with increased levels of nitric oxide (NO). In both inoculated mice with Ltrop or Lmj, parasite load showed a significant decrease at later steps during the CL course post‐treatment. In vivo bioluminescence intensity did not show any ARG's inhibitory effect on treated‐Ltrop. The findings verified that the ARG activity may partially control the L. tropica infection in BALB/c mice through reduction of parasite proliferation and parasite killing through NO generation. This effect is dose‐dependent.