High-throughput CZE-UV determination of arginine and dimethylated arginines in human plasma

Experimental studies document that increased asymmetric dimethylarginine (ADMA) blood levels inhibit NOS significantly, reducing NO generation. ADMA measurement often needs sample cleanup by SPE prior to chromatography and precolumn derivatization that cannot be easily employed in a routine clinical setting. We set up a new reliable CE method to measure ADMA, symmetric dimethylarginine (SDMA), and arginine without sample extraction or precolumn derivatization in order to examine their concentrations in human plasma. Sample was concentrated prior to CE injection and analytes were monitored by UV detection. CE analysis was performed in an uncoated fused-silica capillary, 75 microm id and 60.2 cm length (50 cm to the detection window), injecting 1 s water plug (0.5 psi) followed by 10 s of the sample (0.5 psi). Separation was carried out in a 50 mmol/L Tris-phosphate run buffer at pH 2.30, 15 degrees C and 15 kV (75 microA) at normal polarity. Recovery of plasma ADMA was 101-104% and inter-day CV was less than 3%. Assay performance was evaluated measuring the levels of arginine and its dimethyl derivatives in 77 subjects. Passing-Bablok regression and Bland-Altman test for methods comparison suggest that the data obtained by our method and by a reference CE-LIF assay are similar.