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High performance liquid chromatographic method for the determination of homocysteine in human plasma using a water-soluble phosphine reducing agent and fluorogenic derivatization

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Total homocysteine in human plasma was determined by reducing plasma disulfides to thiols using a water-soluble reducing agent, tris-(carboxyethyl)-phosphine (TCEP) at room temperature for 10 minutes. Reduced plasma samples were derivatized using the thiol-specific fluorogenic reagent 4-(aminosulfonyl)-7-fluoro-2,1,3-benzoxadiazole (ABD-F) in borate borate buffer. The derivatized homocysteine, cysteine, glutathione, and Cys-Gly peptide were separated within 10 minutes using a reserved-phase column with an isocratic elution method, giving detection sensitivity in the low picomole level.

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Authors and Affiliations

  1. Center for Bioanalytical Research, Higuchi Bioscience Center, University of Kansas, 2099 Constant Avenue, 66047, Lawrence, KS, USA

    M. C. Whittle, J. F. Stobaugh & O. S. Wong

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  1. M. C. Whittle
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  2. J. F. Stobaugh
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  3. O. S. Wong
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Whittle, M.C., Stobaugh, J.F. & Wong, O.S. High performance liquid chromatographic method for the determination of homocysteine in human plasma using a water-soluble phosphine reducing agent and fluorogenic derivatization. Chromatographia 52 (Suppl 1), S78–S82 (2000). https://doi.org/10.1007/BF02493130

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  • DOI: https://doi.org/10.1007/BF02493130

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