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Determination of glutathione and glutathione disulfide in human whole blood using HPLC with coulometric detection: a comparison with fluorescence detection
Authors: Kanďár Roman | Žáková Pavla | Marková Miroslava | Lotková Halka | Kučera Otto | Červinková Zuzana
Year: 2011
Type of publication: článek v odborném periodiku
Name of source: Collection of Czechoslovak Chemical Communications
Publisher name: Ústav organické chemie a biochemie AV ČR
Place: Praha
Page from-to: 274-294
Titles:
Language Name Abstract Keywords
cze Stanovení glutathionu a glutathiondisulfidu v lidské plné krvi metodou HPLC s coulometrickou detekcí: porovnání s fluorescenční detekcí Popisujeme zde relativně jednoduchou metodu pro stanovení glutathionu (GSH) a glutathiondisulfidu (GSSG) v lidské plné krvi. Pro simultánní stanovení GSH a GSSG jsme použili HPLC s coulometrickou elektrochemickou detekcí. Naředěné a přefiltrované extrakty byly dávkovány přímo na kolonu Polaris 5u C18-A, 250 x 4,6 mm a isokraticky eluovány. Glutathion v extraktu naředěném kyselinou chlorovodíkovou (1 mmol/l) byl stabilní při 4 °C minimálně 8 hodin. Analytické parametry metody jsou vyhovující: koeficienty přesnosti v sérii a mezi sériemi byly pod 10 %. Výtěžnost metody byla v intervalu 91,6-97,6 % pro GSH a 85,0-104,4 % pro GSSG.Linearita metody byla v rozsahu 5,0-2000 μmol/l (mez detekce 2,1 μmol/l) pro GSH a 2,0-250,0 μmol/l (mez detekce 0,9 μmol/l) pro GSSG. glutathion;glutathiondisulfid;stabilita glutathionu;kapalinová chromatografie;coulometrická elektrochemická a fluorescenční detekce
eng Determination of glutathione and glutathione disulfide in human whole blood using HPLC with coulometric detection: a comparison with fluorescence detection We describe a relatively simple method for the determination of glutathione (GSH) and glutathione disulfide (GSSG) in human whole blood. We have used an HPLC with coulometric electrochemical detection for the simultaneous measurement of GSH and GSSG. Diluted and filtered trichloroacetic acid extracts were injected directly into the HPLC system and were eluted isocratically on a Polaris 5u C18-A, 250 x 4.6 mm analytical column. Glutathione in samples extracted with trichloroacetic acid and diluted with 1.0 mmol/l hydrochloric acid was stable at 4 °C for at least 8 h. The analytical performance of this method is satisfactory: the intra-assay and inter-assay coefficients of variation were below 10%. Quantitative recoveries from spiked whole blood samples were at intervals 91.6-97.6% for GSH and 85.0-104.4% for GSSG. The linear range is 5.0-2000.0 μmol/l, with a detection limit of 2.1 μmol/l (signal-to-noise ratio = 3) for GSH and 2.0-250.0 μmol/l, with a detection limit of 0.9 μmol/l for GSSG. glutathione;glutathione disulfide;glutathione stability;liquid chromatography;coulometric electrochemical and fluorescence detection

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