The simple isocratic HPLC-UV method for the simultaneous determination of reduced and oxidized glutathione in animal tissue

• Autorzy: Begic A. , Djuric A. , Gobeljic B. , Stevanovic I. , Lukic V. , Stanojevic I. , Ninkovic M. , Saso L. , Vojvodic D. , Djukic M.

Identyfikatory

DOI

10.1556/1326.2017.29.1.5

• Języki publikacji: EN

Abstrakty

EN

The aim of our work was to optimize and apply simple high-performance liquid chromatography method with ultraviolet detection (HPLC-UV) for simultaneous determination of reduced (GSH) and oxidized (GSSG) glutathione in biological matrix (specifically, the rat liver tissue was used herein), since the ratio between oxidized and reduced glutathione forms (GSSG-GSH) has been recognized as an important biological marker of oxidatively depleted GSH in oxidative stress (OS)-associated diseases and poisonings. An isocratic chromatographic separation of GSH and GSSG (2.8 min and 6.3 min, respectively) was performed with the mobile phase consisted of sodium perchlorate solution (pH adjusted to 2.8) at flow rate of 1 mL min−1, detection set at 215 nm, and column temperature of 40 °C. The method offers short run time, linearity in the range of 0.01-200 μM concentration for both compounds (R2 = 1), low limits of detection and quantification (GSH: 0.18 μM and 0.56 μM, GSSG: 0.52 μM and 1.58 μM, respectively), precision, accuracy (bias < 2%), and high reproducibility. Through suitable sample handling, an overestimation of GSSG was prevented. High recovery (>99%) was achieved. The method was successfully applied for the analysis of GSH and GSSG in liver homogenates of Wistar rats intraperitoneally exposed to cadmium (Cd) (1 mg kg−1 CdCl2/21 days). Regardless of other Cd-mediated hepatotoxicity mechanisms, herein, we have exclusively interpreted/emphasized oxidative GSH depletion. The presented method is acceptable for a routine analysis of GSH and GSSG in biological matrix, while the calculated ratio GSSG-GSH is considered as a valuable OS marker.

Słowa kluczowe

EN

oxidized/reduced glutathione; HPLC-UV; cadmium; hepatotoxicity

• Wydawca: University of Silesia in Katowice ; Akademiai Kiado
• Czasopismo: Acta Chromatographica
• Rocznik: 2017
• Tom: Vol. 29, no. 1
• Strony: 67--84
• Opis fizyczny: Bibliogr. 21 poz., rys.

Twórcy

autor

Begic A.

• Department for Toxicology, Faculty of Pharmacy, University of Belgrade, Vojvode Stepe 450, 11221 Belgrade, Serbia

autor

Djuric A.

• Department for Toxicology, Faculty of Pharmacy, University of Belgrade, Vojvode Stepe 450, 11221 Belgrade, Serbia

autor

Gobeljic B.

• Department for Toxicology, Faculty of Pharmacy, University of Belgrade, Vojvode Stepe 450, 11221 Belgrade, Serbia

autor

Stevanovic I.

• Institute for Medical Research of Medical Military Academy, Crnotravska 17, 11000 Belgrade, Serbia

autor

Lukic V.

• Institute of Forensic Medicine, Faculty of Medicine, University of Belgrade, 31a Deligradska Street, 11000 Belgrade, Serbia

autor

Stanojevic I.

• Institute for Medical Research of Medical Military Academy, Crnotravska 17, 11000 Belgrade, Serbia

autor

Ninkovic M.

• Institute for Medical Research of Medical Military Academy, Crnotravska 17, 11000 Belgrade, Serbia

autor

Saso L.

• Department of Physiology and Pharmacology, Sapienza University, Rome, Italy

autor

Vojvodic D.

• Institute for Medical Research of Medical Military Academy, Crnotravska 17, 11000 Belgrade, Serbia

autor

Djukic M.

• Department for Toxicology, Faculty of Pharmacy, University of Belgrade, Vojvode Stepe 450, 11221 Belgrade, Serbia

Bibliografia

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Uwagi

PL

Opracowanie ze środków MNiSW w ramach umowy 812/P-DUN/2016 na działalność upowszechniającą naukę (zadania 2017).