ADMA levels
Blood samples were collected in tubes containing EDTA. After centrifugation, plasma was immediately collected and stored at −80 °C until processing for analysis. Levels of ADMA, SDMA, and arginine were measured by high performance liquid chromatography (HPLC) with fluorescence detection and using monomethyl-arginine as internal standard. Plasma samples were separated using solid-phase extraction, and analytes were derivatized with ortho-phthalaldehyde containing 3-mercaptopropionic acid, as previously described [24]. Methylarginines levels were also measured in plasma obtained from participants in clinical trial NCT00607672 of subjects undergoing cardiac surgery, who were randomized to one week treatment with ramipril, candesartan or placebo prior to surgery.