DEVELOPMENT OF METHODIC OF TOTAL GLUTATHIONE AMOUNT DETERMINATION
The result's identifiers
Result code in 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>
Result on the web
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DOI - Digital Object Identifier
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Alternative languages
Result language
angličtina
Original language name
DEVELOPMENT OF METHODIC OF TOTAL GLUTATHIONE AMOUNT DETERMINATION
Original language description
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.
Czech name
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Czech description
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Classification
Type
D - Article in proceedings
CEP classification
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OECD FORD branch
10406 - Analytical chemistry
Result continuities
Project
Result was created during the realization of more than one project. More information in the Projects tab.
Continuities
P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)
Others
Publication year
2012
Confidentiality
S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů
Data specific for result type
Article name in the collection
MENDELNET 2012
ISBN
978-80-7375-836-3
ISSN
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e-ISSN
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Number of pages
7
Pages from-to
1218-1224
Publisher name
Neuveden
Place of publication
Neuveden
Event location
Mendel Univ, Fac Agron, Brno
Event date
Nov 21, 2012
Type of event by nationality
CST - Celostátní akce
UT code for WoS article
000366461200142