Determination of L-arginine and NG,NG- and NG,NG′-dimethyl-L-arginine in plasma by liquid chromatography as AccQ-Fluor™ fluorescent derivatives

A new HPLC assay for the detection Of L-arginine, N-G,N-G-dimethyl-L-arginine (ADMA) and N-G,N-G'-dimethyl-L-arginine (SDMA) in plasma using the derivatisation reagent AccQ-Fluor(TM) (6-aminoquinolyl-N-hydroxysuccinimidyl carbamate) is described. The fluorescent derivatives produced are extremely stable enabling routine processing of large numbers of samples. Arginine and its metabolites are extracted from plasma on strong cation exchange (SCX) cartridges with N-G-monomethyl-L-arginine (NMMA) as internal standard, derivatised and separated on a C 18 column with acetonitrile in 0.1 M sodium acetate buffer pH 6. Separation of the stereoisomers ADMA and SDMA was excellent and improvements to the solid phase extraction (SPE) procedure enabled good recovery (>80%) of arginine, ADMA and SDMA. The utility of the method is exemplified by comparison of plasma concentrations of ADMA, SDMA and arginine in healthy volunteers and diabetic/ischaemic patients. (C) 2004 Elsevier B.V. All rights reserved.