Effects of PDE5 inhibitors on vasculogenic ED Savas M et al. deposition.11 The negative effect of free radicals is mediated by degradative agents such as proteolytic enzymes and the final step of collagen degradation is mediated by proli- dase.12 Surazynski et al. noted that prolidase may also have a role in angiogenesis since prolidase deficiency is asso- ciated with angiopathy.13 In this study, we investigated the acute effects of PD5 inhibitor (tadalafil citrate 20 mg) as an on-demand use PDE5 inhibitor at peak serum level on serum oxidative status and prolidase enzyme activities in patients with ED. MATERIAL AND METHODS Subjects The study included 36 patients with clinically documented ED (aged 37–59 years, mean¡SD 49.5¡6.5) who had been followed up at the department of urology, had a stable monogamous relationship with a female partner, and for whom PDE5 inhibitor treatment had been recently proposed. Controls comprised 30 people with no ED (aged 35–58 years, mean¡SD 50.1¡6.7). The study was approved by Harran University’s Institutional Review Board and was performed in accordance with the ethical standards laid down in the 1975 and 1983 Declaration of Helsinki. The participants were informed about the study protocol and written consent was obtained from all participants before the start of the study. All patients provided a detailed sexual history, and underwent a physical examination, blood chemistry and endocrine assay, and color Doppler ultrasonography during pharmacologically induced and sexually stimulated erec- tion. Patients with angina during intercourse, unstable angina or any other evidence of recently diagnosed coronary artery disease, poorly controlled blood pressure or orthostatic hypotension, congestive heart failure, arrhyth- mia, significant renal or hepatic dysfunction, anemia, and patients aged .60 years were excluded. Additionally, patients who had been receiving any medication that had an effect on serum oxidative status (vitamin E, vitamin C, L- arginine, etc) within the past 4 weeks or who had under- gone pelvic surgery were also excluded. All blood samples for determination of total antioxidant status (TAS), total oxidant status (TOS), and prolidase activities were collected from the subjects just before and 2 h after 20 mg oral tadalafil citrate (CialisH, Lilly Inc, Istanbul, Turkey) admin- istration—that is, at the peak plasma level, which occurs at approximately 2 h after administration. During these 2 h, subjects remained at rest under clinic conditions and were not allowed to consume anything except water. Blood sample collections were performed in an air-conditioned consulting room at a temperature of 23–25˚C. Measurement Total Oxidant–Antioxidant Status An automated colorimetric measurement method was used for the analysis of TOS.14 The results were expressed as mmol H2O2 Eq/L. TAS was measured by an automated measurement method in which hydroxyl radicals are produced via the Fenton reaction and the colored dianisi- dinyl radical cations consequently produced in the reaction medium of the assay can be measured by absorbance.15 The results were expressed as mmol Trolox Eq/L. Measurement of Prolidase Activity Serum was diluted 40-fold with 25 mmol/L Mn2+, 40 mmol/L Trizma HCl buffer (pH 8.0) and preincubated CLINICS 2010;65(12):1311-1314 at 37˚C for 2 h. The reaction mixture containing 30 mmol/L Gly-Pro, 40 mmol/L Trizma HCl buffer (pH 8.0), and 100 mL of preincubation serum in 1 mL was incubated at 37˚C for 30 min. The reaction was stopped by adding 0.5 mL 20% trichloroacetic acid solution. The supernatant was used for measurement of proline by the method proposed by Myara et al.,16 which is a modification of Chinard’s method.17 Statistical Analysis Statistical analyses were performed using the Statistical Package for the Social Sciences (SPSS Inc, Chicago, IL, USA) version 11.0 for Windows. Results were expressed as mean¡SD for all continuous variables. Differences between control, pre- and post-medication groups were assessed using an independent samples t-test. Differences between pre- and post-medication groups were assessed using a paired sample t-test. A value of p,0.05 was considered statistically significant. RESULTS Demographic characteristics and risk factors for ED of the patients are given in Table 1. The serum samples of 30 control cases and 36 patients with ED (body mass index; range 22.0– 29.8 kg/m 2 , mean¡SD 27.0¡2.2 kg/m2), before and after tadalafil citrate administration (120 min), were analyzed. For controls, serum levels of TOS, TAS, and prolidase were 4.37¡0.9 mmol H2O2 Eq/L, 1.83¡0.2 mmol Trolox Eq/L, and 218.09¡17.5 U/L, respectively. The mean basal and post tadalafil citrate administration serum parameters (TOS, TAS, and prolidase) of the patients are summarized in Table 2. While the mean basal serum level of TAS and TOS were 1.1¡0.0 and 10.3¡1.1 mmol H2O2 Eq/L, the post tadalafil citrate values were 1.6¡0.0 and 6.9¡1.2 mmol Trolox Eq/L, respectively. Both reductions in TOS and increases in TAS serum values were significant (p,0.0001). The mean proli- dase activity under the influence of the drug was 228.2¡ 19.2 U/L, in comparison with the basal value of 236.4¡ 19.5 U/L—a significant reduction (p,0.0001). DISCUSSION This study aimed to document the acute effects of tada- lafil citrate at peak serum levels on the vascular system by evaluating serum oxidative–antioxidative status, and proli- dase enzyme activities. Tadalafil citrate was chosen for this study, because it has the longest half-life of the PDE5 Table 1 - Demographic characteristics and risk factors for erectile dysfunction of the patients and control cases. Control (n = 30) Patients (n = 36) p Value 1312 Demographic characteristics Age (years) (mean ¡SD) BMI (kg/m2) Risk factors n (%) Smoking Depression Diabetes mellitus Hypertension Chronic heart failure Hypercholesterolemia 50.1¡6.7 28.0¡2.9 12 (40) - 8 (27) 13 (43) 5 (17) 4 (13) BMI, body mass index. 49.5¡6.5 27.0¡2.2 14 (39) 2 (6) 12 (33) 12 (33) 7 (19) 7 (19) .0.05 .0.05