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DEVELOPMENT OF METHODIC OF TOTAL GLUTATHIONE AMOUNT DETERMINATION

Identifikátory výsledku

  • Kód výsledku v IS VaVaI

    <a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00216305%3A26620%2F12%3APU126848" target="_blank" >RIV/00216305:26620/12:PU126848 - isvavai.cz</a>

  • Výsledek na webu

  • DOI - Digital Object Identifier

Alternativní jazyky

  • Jazyk výsledku

    angličtina

  • Název v původním jazyce

    DEVELOPMENT OF METHODIC OF TOTAL GLUTATHIONE AMOUNT DETERMINATION

  • Popis výsledku v původním jazyce

    Glutathione plays important role in biochemical processes associated with oxidative stress in bacterial, plant and animals cells. Glutathione is occurring in the reduced (GSH) and oxidized form (GSSG) in the organism. The ratio between GSH and GSSG is considered as a marker of oxidative stress. During the sample preparation the destabilization of the GSH and GSSG forms is occurring which might be a problem for subsequent analysis. Aim of this study was the optimization and of quantitative reduction GSSG to GSH for HPLC-ED determination using tris (2-carboxyethyl) phosphin (TCEP) and ethylendiamintetracetic acid (EDTA). The efficiency of reduction of GSSG was analyzed using high pressure liquid chromatography with electrochemical detection (HPLC-ED). For GSH and GSSG determination we used gradient elution with methanol on reversed-phase column and coulometric detection. Limit of detection for GSH and GSSG was determined as 30 nM and 200 nM respectively. For qualitative reduction of GSSG to GSH we optimized the concentration of reducing agent (1, 2, 4, 6 mM), time of reaction (1, 5, 10, 15, 20, 25, 30, 40, 45, 50, 55, 60 minutes) and pH range of tested buffers (Britton-Robinson, Borate buffer, Phosphate buffer and Acetate buffer). The influence of addition of EDTA was also tested. We determined the optimal condition of the reducing reaction as 2 mM TCEP in Borate buffer (pH 9) and reaction time 15 minutes. No influence of EDTA addition on the reducing reaction was observed. The usage of the method was estimated for concentration range 60 nM - 200 mu M of GSSG.

  • Název v anglickém jazyce

    DEVELOPMENT OF METHODIC OF TOTAL GLUTATHIONE AMOUNT DETERMINATION

  • Popis výsledku anglicky

    Glutathione plays important role in biochemical processes associated with oxidative stress in bacterial, plant and animals cells. Glutathione is occurring in the reduced (GSH) and oxidized form (GSSG) in the organism. The ratio between GSH and GSSG is considered as a marker of oxidative stress. During the sample preparation the destabilization of the GSH and GSSG forms is occurring which might be a problem for subsequent analysis. Aim of this study was the optimization and of quantitative reduction GSSG to GSH for HPLC-ED determination using tris (2-carboxyethyl) phosphin (TCEP) and ethylendiamintetracetic acid (EDTA). The efficiency of reduction of GSSG was analyzed using high pressure liquid chromatography with electrochemical detection (HPLC-ED). For GSH and GSSG determination we used gradient elution with methanol on reversed-phase column and coulometric detection. Limit of detection for GSH and GSSG was determined as 30 nM and 200 nM respectively. For qualitative reduction of GSSG to GSH we optimized the concentration of reducing agent (1, 2, 4, 6 mM), time of reaction (1, 5, 10, 15, 20, 25, 30, 40, 45, 50, 55, 60 minutes) and pH range of tested buffers (Britton-Robinson, Borate buffer, Phosphate buffer and Acetate buffer). The influence of addition of EDTA was also tested. We determined the optimal condition of the reducing reaction as 2 mM TCEP in Borate buffer (pH 9) and reaction time 15 minutes. No influence of EDTA addition on the reducing reaction was observed. The usage of the method was estimated for concentration range 60 nM - 200 mu M of GSSG.

Klasifikace

  • Druh

    D - Stať ve sborníku

  • CEP obor

  • OECD FORD obor

    10406 - Analytical chemistry

Návaznosti výsledku

  • Projekt

    Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

  • Návaznosti

    P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Ostatní

  • Rok uplatnění

    2012

  • Kód důvěrnosti údajů

    S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Údaje specifické pro druh výsledku

  • Název statě ve sborníku

    MENDELNET 2012

  • ISBN

    978-80-7375-836-3

  • ISSN

  • e-ISSN

  • Počet stran výsledku

    7

  • Strana od-do

    1218-1224

  • Název nakladatele

    Neuveden

  • Místo vydání

    Neuveden

  • Místo konání akce

    Mendel Univ, Fac Agron, Brno

  • Datum konání akce

    21. 11. 2012

  • Typ akce podle státní příslušnosti

    CST - Celostátní akce

  • Kód UT WoS článku

    000366461200142