The aim of the present study was to investigate serum homocysteine levels in patients with erectile dysfunction and to evaluate the relationship between serum homocysteine levels and response to the standard 50 mg phosphodiesterase 5 inhibitor treatment. Twenty-eight erectile dysfunction patients having normal vascular parameter according to Penile Doppler Ultrasonography and twenty healthy subjects were enrolled in the study. All subjects filled The International Index of Erectile Function (IIEF) questionnaire. A total of 4-6 doses of phosphodiesterase 5 inhibitor (sildenafil 50 mg) were given to patients. Later, they were divided into two groups as sildenafil responder and non-responder. Serum homocysteine levels were compared in groups based on sildenafil response: Compared with healthy subject, higher homocysteine levels were observed in patients with erectile dysfunction (p = 0.005), especially in sildenafil non-responder group (p = 0.005). There was significant negative correlation between homocysteine and TIFF scores in group responder to sildenafil treatment (r = -0.698, p = 0.008). Mean IIEF scores of patients with non-responder to sildenafil 50 mg were lower than those of controls (p = 0.0001), but mean IIEF scores of patients with responders approached values observed in control subjects (p = 0.002). The results indicated that measurement of serum homocysteine levels could be used as a marker for the evaluation of efficacy of phosphodiesterase 5 inhibitor and the selection of efficacious alternative therapies.