HPLC analysis of asymmetric dimethylarginine, symmetric dimethylarginine, homoarginine and arginine in small plasma volumes using a Gemini-NX column at high pH

Catherine Jones, Christabelle Darcy, Tonia Woodberry, Nicholas Anstey, Yvette McNeill

    Research output: Contribution to journalArticle

    29 Citations (Scopus)

    Abstract

    There is increasing recognition of the clinical importance of endogenous nitric oxide synthase inhibitors in critical illness. This has highlighted the need for an accurate high performance liquid chromatography (HPLC) method for detection of asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA) in small volumes of blood. Here, the validation of an accurate, precise HPLC method for the determination of ADMA, SDMA, homoarginine and arginine concentrations in plasma is described. Solid phase extraction is followed by derivatisation with AccQ-Fluor™ and reversed phase separation on a Gemini-NX column at pH 9. Simultaneous detection by both UV-vis and fluorescence detectors affords extra validation. This solid phase extraction method gives absolute recoveries of more than 85% for ADMA and SDMA and relative recoveries of 102% for ADMA and 101% for SDMA. The intra-assay relative standard deviations are 2.1% and 2.3% for ADMA and SDMA, respectively, with inter-assay relative standard deviations of 2.7% and 3.1%, respectively. Advantages of this method include improved recovery of all analytes using isopropanol in the solid phase extraction; sharp, well-resolved chromatographic peaks using a high pH mobile phase; a non-endogenous internal standard, n-propyl l-arginine; and accurate and precise determination of methylated arginine concentrations from only 100 μL of plasma.

    Original languageEnglish
    Pages (from-to)8-12
    Number of pages5
    JournalJournal of Chromatography B-Analytical Technologies in The Biomedical and Life Sciences
    Volume878
    Issue number1
    DOIs
    Publication statusPublished - 1 Jan 2009

    Keywords

    • Arginine
    • Asymmetric dimethylarginine
    • High performance liquid chromatography
    • Homoarginine
    • Symmetric dimethylarginine

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