To evaluate the relationship between serum endothelial cell specific molecule-1 (Endocan), asymmetric dimethylarginine (ADMA) values, and penile Doppler ultrasonography (USG) findings in patients with severe erectile dysfunction (ED). This prospective study included 73 patients who were classified as severe ED and had an indication for penile Doppler USG in our urology outpatient clinic between April 2017 and January 2020. Fasting blood sugar, lipid profile, thyroid function tests, total testosterone, and serum Endocan and ADMA values were sampled, and penile Doppler USG examination data were recorded. Vasculogenic ED was detected in 51 (69.86%) of 73 patients, while the flow rates were normal in 22 (30.14%). Among those with vasculogenic ED, 15 (29.41%) had arterial insufficiency, 22 (43.13%) had venous leakage, and 14 (27.46%) had mixed. There was no statistically significant difference between the mean ADMA and Endocan values of ED with normal flow (14.44 ± 6.20 ng/mL and 0.18 ± 0.14 ng/mL) and the vasculogenic ED (12.31 ± 5.86 ng/mL and 0.21 ± 0.16 ng/mL) groups (P = .097 for ADMA and P = .315 for Endocan, respectively). Endocan and ADMA levels were not predictors of severe ED. There is a need for multicenter studies with larger patient populations to be conducted with different biomarkers that may have a higher predictive value in the future.