Simultaneous determination of multiple amino acids in plasma in critical illness by high performance liquid chromatography with ultraviolet and fluorescence detection

Hao Wang, Yvette McNeil, Tsin Yeo, Nicholas Anstey

    Research output: Contribution to journalArticleResearchpeer-review

    Abstract

    There is increasing recognition that the host response to critical illness includes derangement of multiple amino acid pathways, including amino acids (AAs) central to metabolism and immune, endothelial and neurological function. To characterise concentration changes of these plasma amino acid we report the development and validation of a method for the quantification of AAs in small volumes of plasma (50. ?L) using HPLC with simultaneous UV and fluorescence (FL) detection. Protein precipitation and pre-column derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) is followed by reversed phase HPLC separation. Calibration curves were built with norleucine as an internal standard. Thirty-three (including the 20 proteinogenic) AAs, were selected as standards and their corresponding concentrations in the plasma of healthy human controls and patients with severe falciparum malaria were quantified. This method enables the detection of perturbations in arginine metabolism, aromatic amino acid pathways, methionine transsulfuration and transmethylation pathways and other metabolic pathways. � 2013 Elsevier B.V.
    Original languageEnglish
    Pages (from-to)53-58
    Number of pages6
    JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
    Volume940
    DOIs
    Publication statusPublished - 2013

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    High performance liquid chromatography
    Critical Illness
    Fluorescence
    High Pressure Liquid Chromatography
    Plasmas
    Amino Acids
    Metabolism
    Norleucine
    Aromatic Amino Acids
    Falciparum Malaria
    Plasma Volume
    Metabolic Networks and Pathways
    Phase separation
    Methionine
    Calibration
    Arginine
    Proteins

    Cite this

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    abstract = "There is increasing recognition that the host response to critical illness includes derangement of multiple amino acid pathways, including amino acids (AAs) central to metabolism and immune, endothelial and neurological function. To characterise concentration changes of these plasma amino acid we report the development and validation of a method for the quantification of AAs in small volumes of plasma (50. ?L) using HPLC with simultaneous UV and fluorescence (FL) detection. Protein precipitation and pre-column derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) is followed by reversed phase HPLC separation. Calibration curves were built with norleucine as an internal standard. Thirty-three (including the 20 proteinogenic) AAs, were selected as standards and their corresponding concentrations in the plasma of healthy human controls and patients with severe falciparum malaria were quantified. This method enables the detection of perturbations in arginine metabolism, aromatic amino acid pathways, methionine transsulfuration and transmethylation pathways and other metabolic pathways. � 2013 Elsevier B.V.",
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    T1 - Simultaneous determination of multiple amino acids in plasma in critical illness by high performance liquid chromatography with ultraviolet and fluorescence detection

    AU - Wang, Hao

    AU - McNeil, Yvette

    AU - Yeo, Tsin

    AU - Anstey, Nicholas

    PY - 2013

    Y1 - 2013

    N2 - There is increasing recognition that the host response to critical illness includes derangement of multiple amino acid pathways, including amino acids (AAs) central to metabolism and immune, endothelial and neurological function. To characterise concentration changes of these plasma amino acid we report the development and validation of a method for the quantification of AAs in small volumes of plasma (50. ?L) using HPLC with simultaneous UV and fluorescence (FL) detection. Protein precipitation and pre-column derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) is followed by reversed phase HPLC separation. Calibration curves were built with norleucine as an internal standard. Thirty-three (including the 20 proteinogenic) AAs, were selected as standards and their corresponding concentrations in the plasma of healthy human controls and patients with severe falciparum malaria were quantified. This method enables the detection of perturbations in arginine metabolism, aromatic amino acid pathways, methionine transsulfuration and transmethylation pathways and other metabolic pathways. � 2013 Elsevier B.V.

    AB - There is increasing recognition that the host response to critical illness includes derangement of multiple amino acid pathways, including amino acids (AAs) central to metabolism and immune, endothelial and neurological function. To characterise concentration changes of these plasma amino acid we report the development and validation of a method for the quantification of AAs in small volumes of plasma (50. ?L) using HPLC with simultaneous UV and fluorescence (FL) detection. Protein precipitation and pre-column derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) is followed by reversed phase HPLC separation. Calibration curves were built with norleucine as an internal standard. Thirty-three (including the 20 proteinogenic) AAs, were selected as standards and their corresponding concentrations in the plasma of healthy human controls and patients with severe falciparum malaria were quantified. This method enables the detection of perturbations in arginine metabolism, aromatic amino acid pathways, methionine transsulfuration and transmethylation pathways and other metabolic pathways. � 2013 Elsevier B.V.

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    KW - Fluorescence detection

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    KW - Simultaneous determinations

    KW - High performance liquid chromatography

    KW - Metabolism

    KW - Physiology

    KW - Plasmas

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    KW - 4 hydroxy proline

    KW - 6 aminoquinolyl n hydroxysuccinimidyl carbamate

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    KW - aspartic acid

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    KW - cystathionine

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    KW - diaminobutyric acid

    KW - glutamic acid

    KW - glutamine

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    KW - human tissue

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    KW - limit of quantitation

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    KW - Adult

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    KW - Reproducibility of Results

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    KW - Sepsis

    KW - Spectrometry, Fluorescence

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    U2 - 10.1016/j.jchromb.2013.09.016

    DO - 10.1016/j.jchromb.2013.09.016

    M3 - Article

    VL - 940

    SP - 53

    EP - 58

    JO - Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences

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    SN - 1570-0232

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