Rapid simultaneous determination of arginine and methylated arginines in human urine by high-performance liquid chromatography–mass spectrometry
ABSTRACT A simple, sensitive and fast method using reversed-phase high-performance liquid chromatography (HPLC)–mass spectrometry (MS) coupling with an atmospheric pressure chemical ionization (APCI) interface was developed for simultaneous separation and determination of l-arginine (ARG), NG,NG-dimethylarginine (ADMA) and NG,N′G-dimethylarginine (SDMA) in human urine. This method involved the use of the [M+H]+ ions of ARG, ADMA and SDMA at m/z 175, 203 and 203 in the selective ion monitoring (SIM) mode. Satisfactory separation was achieved on a mm Shimadzu VP-ODS column by using the mobile phase consisting of water (95%), acetonitrile (5%) and trifluoroacetic acid (TFA, 0.4%). l-Homoarginine was used as the internal standard for the assay. With an isocratic HPLC, the total LC–APCI–MS analysis time was less than 5 min, making the method the fastest and most specific method reported to date. The limits of quantification (LOQ) were found to be 0.2 μmol l−1 for ARG, ADMA and SDMA. The inter-assay precision and accuracy were in the range of 2.1–6.8 and −4.4–5.4%, respectively. The intra-assay precision and accuracy were in the order of 1.6–4.1 and −1.8–3.2%, respectively. The recoveries were between 98.2 and 103.2%. With the help of the proposed method, the levels of ARG, ADMA and SDMA in human urine were determined.