Simultaneous determination of l-citrulline and l-arginine in plasma by high performance liquid chromatography

ObjectiveTo develop a method for simultaneously determining l-citrulline and l-arginine levels in plasma using RP-HPLC with ultraviolet detection.Design and methodsPlasma samples were deproteinized by trichloroacetic acid and heat. Phenyl-isothiocyanate (PITC) solution was used as derivatization reagent and a gradient elution was carried out.ResultsThe linearity for l-citrulline and l-arginine ranged from 0 to at least 1000 μmol/L. R2 values were above 0.9999 for both. LODs for l-citrulline and l-arginine were 0.0201 μmol/L and 0.0476 μmol/L, respectively, while LOQs were 0.240 μmol/L and 0.448 μmol/L, respectively. Intra- and inter-day CVs were less than 3.40% and 7.2%, respectively. The average recovery was from 86.22% to 118.9%. l-Citrulline and l-arginine concentrations in healthy controls were 60.77 ± 9.18 μmol/L and 58.19 ± 16.43 μmol/L, respectively.ConclusionThis approach offers a reliable, efficient analytical platform for the simultaneous determination of citrulline and arginine levels in plasma.