Improved method for simultaneous determination of L-arginine and its mono- and dimethylated metabolites in biological samples by high-performance liquid chromatography

An improved method has been developed for the determination of L-arginine and its methylated metabolites, N-G-monomethyl-L-arginine (L-NMMA), N-G,N-G-dimethyl-L-arginine (asymmetric DMA, ADMA) and N-G,N-G'-dimethyl-L-arginine (symmetric DMA, SDMA) in biological samples. Extraction of these compounds with a strong cation-exchange resin AG50W-X8 with L-homoarginine (2-amino-6-guanidinohexanoic acid) as an internal standard gave a recovery of more than 70% except for SDMA from plasma samples. After extracted samples were converted to fluorescent derivatives with o-phthalaldehyde (OPA) in an alkaline medium, the following high-performance liquid chromatographic separation with a ODS column (wide-pore size, 300 Angstrom) was successfully performed with an isocratic mobile phase system. The method permits quantitative determination of L-arginine and its methylated metabolites at concentrations as low as 4 mu M and 0.18 mu M, respectively. Using this method, the levels of L-arginine, L-NMMA, ADMA and SDMA in human plasma, urine and rat tissue were determined. (C) 2000 Elsevier Science B.V. All rights reserved.