A new derivatization method coupled with LC-MS/MS to enable baseline separation and quantification of dimethylarginines in human plasma from patients to receive on-pump CABG surgery

{Asymmetric dimethylarginine (ADMA) is an inhibitor of nitric oxide synthase and a risk factor for cardiovascular events. We have developed a new derivatization method to enable baseline separation of the regio-isomers, ADMA, and symmetric dimethylarginine (SDMA), within 15 min on a C18 reverse phase column. Reacting naphthalene-2,3-dicarboxaldehyde with ADMA and SDMA in the presence of 2-mercaptoethanol produces corresponding 2,3-dihydro-benzo{[}f]isoindol-1-ones that are more stable than previously reported ortho-phthaldialdehyde and 2-mercaptoethanol derivatives. LC-MS/MS quantitation of these derivatives can be used to determine ADMA and SDMA concentrations in the plasma of patients to receive on-pump coronary artery bypass grafting (CABG) surgery. The LOD, LOQ and lower LOQ (LLOQ) of this method were determined to be 2.6, 8.7, and 25 nM for ADMA, and 2.5, 8.3, and 25 nM for SDMA, respectively, with consumption of only 50 mu L of plasma. The relative standard deviations and relative errors of the intraday and interday determinations, as measurements of reproducibility and accuracy, are all within 15%. The ADMA and SDMA concentrations in patient plasma are 298.1 +/- 11.2 nM (mean +/- S.E.M.