Arginase 1 (ARG1) and arginase 2 (ARG2) compete with nitric oxide synthases for the substrate l-arginine. Here we aim to assess whether arginase 1 and 2 plasma levels, plasma arginase activity or genetic factors are associated with altered responsiveness to sildenafil. We studied 71 post-prostatectomy erectile dysfunction (ED) patients (PED group) and 72 clinical ED patients (CED). Patients responded to the International Index of Erectile Function questionnaire before and after the treatment. We found positive and negative correlations between plasma levels of arginase 1 and sildenafil responsiveness in the PED and CED groups, respectively. PED group also presented negative correlation between plasma arginase activity and sildenafil responsiveness. Sildenafil poor responders have shown higher plasma arginase activity in PED and higher arginase 1 levels on CED groups. In addition, variant genotypes for the rs2781659, rs2781667 and rs17599586 polymorphisms were associated with reduced arginase activity, as well as the GTTT ARG1 haplotype in CED group.