Glutathione (GSH) Determination by a Very Simple Electrochemical Method

  • Mustafa Kemal Sezgintürk
  • Erhan Dinçkaya


Glutathione (GSH; gamma-glutamylcysteinylglycine) is ubiquitous in mammalian and other living cells. It contains an unusual peptide linkage between the amine group of cysteine and the carboxyl group of the glutamate side chain. It has several important functions, including protection against oxidative stress. It is synthesized from its constituent amino acids by the consecutive actions of gamma-glutamylcysteine synthetase and GSH synthetase. Cellular levels of GSH may be increased by supplying substrates and GSH delivery compounds. Increasing cellular GSH may be therapeutically useful. In this study, we investigated the applicability of the glassy carbon electrode coated with thin Hg film layer to the determination of reduced glutathione (GSH). For this purpose, firstly, Hg coating process parameters were studied such as concentration of mercury coating solution, coating current, coating temperature. Then, working conditions were investigated. At the end of these studies, we concluded that although some of limitations, the sensor would be applicable to the determination of reduced glutathione.


Glutathione Thin Hg films GSH Biosensor Mercury film electrode 


  1. Álvarez R, Hernández LE, Abadía J, Álvarez-Fernández A (2006) Direct and simultaneous determination of reduced and oxidized glutathione and homoglutathione by liquid chromatography–electrospray/mass spectrometry in plant tissue extracts. Anal Biochem 356:254–264CrossRefGoogle Scholar
  2. Balendiran GK, Dabur R, Fraser D (2004) The role of glutathione in cancer. Cell Biochem Funct 22(6):343–352PubMedCrossRefGoogle Scholar
  3. Brainina KhZ, Malakhova NA, Yu N (2000) Stripping voltammetry in environmental and food analysis. F J Anal Chem 368:307–325CrossRefGoogle Scholar
  4. Cataldi TR, Nardiello D (2005) A pulsed potential waveform displaying enhanced detection capabilities towards sulfur-containing compounds at a gold working electrode. J Chromatogr A 1066:133–142PubMedCrossRefGoogle Scholar
  5. Chow HH, Hakim IA, Vining DR, Crowell JA, Tome ME, Moore JR, Cordova CA, Mikhael DM, Briehl MM, Alberts DS (2007) Modulation of human glutathione S-transferases by polyphenon E intervention. Cancer Epidemiol Biomarkers Prev 16(8):1662–1666PubMedCrossRefGoogle Scholar
  6. Dalton TP, Shertzer HG, Puga A (1999) A regulation of gene expression by reactive oxygen. Ann Rev Pharm Toxic 39:67–101CrossRefGoogle Scholar
  7. Economou A, Fielden PR (1993) Square wave adsorptive stripping voltammetry on mercury film electrodes. Anal Chim Acta 273(1–2):27–34CrossRefGoogle Scholar
  8. Fariss MW, Reed DJ (1987) High-performance liquid chromatography of thiols and disulfides: dinitrophenol derivates. Methods Enzymol 143:101–109PubMedCrossRefGoogle Scholar
  9. Florence TM (1970) Anodic stripping voltammetry with a glassy carbon electrode mercury-plated in situ. J Electroanal Chem 27(2):273–281CrossRefGoogle Scholar
  10. Han YH, Park WH (2009) The effects of N-acetyl cysteine, buthionine sulfoximine, diethyldithiocarbamate or 3-amino-1, 2, 4-triazole on antimycin A-treated Calu-6 lung cells in relation to cell growth, reactive oxygen species and glutathione. Oncol Rep 22(2):385–391PubMedGoogle Scholar
  11. Jagner D, Renman L, Stefandottir SH (1993) Determination of iron(III) and titanium(IV) as their solochrome violet RS complexes by constant-current stripping potentiometry: part 1. Automated single-point calibration method for iron(III). Anal Chim Acta 281(2):305–314CrossRefGoogle Scholar
  12. Kennett EC, Bubb WA, Bansal P, Alewood P, Kuchel PW (2005) NMR studies of exchange between intra- and extracellular glutathione in human erythrocytes. Redox Rep 10:83–90PubMedCrossRefGoogle Scholar
  13. Lakritz J, Plopper CG, Buckpitt AR (1997) Validated high-performance liquid chromatography-electrochemical method for determination of glutathione and glutathione disulfide in small tissue samples. Anal Biochem 247:63–68PubMedCrossRefGoogle Scholar
  14. Liu RM, Vasiliou V, Zhu H, Duh JL, Tabor MW, Puga A, Nebert DW, Sainsbury M, Shertzer HG (1994) Regulation of [Ah] gene battery enzymes and glutathione levels by 5, 10-dihydroindeno[1, 2-b]indole in mouse hepatoma cell lines. Carcinogenesis 15:2347–2352PubMedCrossRefGoogle Scholar
  15. Lomaestro BM, Malone M (1995) Glutathione in health and disease: pharmacotherapeutic issues. Ann Pharmacother 29:1263–1273PubMedGoogle Scholar
  16. Marzal PC, Chumbimuni-Torres KY, Höehr NF, Kubota LT (2006) Determination of glutathione in hemolysed erythrocyte with amperometric sensor based on TTF-TCNQ. Clin Chim Acta 371:152–158CrossRefGoogle Scholar
  17. Meyer AJ, May MJ, Fricker M (2001) Quantitative in vivo measurement of glutathione in Arabidopsis cells. Plant J 27(1):67–78PubMedCrossRefGoogle Scholar
  18. Meyer AJ, Brach T, Marty L, Kreye S, Rouhier N, Jacquot JP, Hell R (2007) Redox-sensitive GFP in Arabidopsis thaliana is a quantitative biosensor for the redox potential of the cellular glutathione redox buffer. Plant J 52(5):973–986PubMedCrossRefGoogle Scholar
  19. New LS, Chan EC (2008) Evaluation of BEH C18, BEH HILIC and HSS T3 (C18) column chemistries for the UPLC/MS/MS analysis of glutathione, glutathione disulfide and ophthalmic acid in mouse liver and human plasma. J Chromatogr Sci 46:209–214PubMedGoogle Scholar
  20. Parmentier C, Leroy P, Wellman M, Nicolas A (1998) Determination of cellular thiols and glutathione-related enzyme activities: versatility of high-performance liquid chromatography–spectrofluorimetric detection. J Chrom B Biomed Sci App 719(1–2):37–46CrossRefGoogle Scholar
  21. Pastore A, Piemonte F, Locatelli M, Lo Russo A, Gaeta LM, Tozzi G, Federici G (2003) Determination of blood total, reduced, and oxidized glutathione in pediatric subjects. Clin Chem 47(8):1467–1469Google Scholar
  22. Patterson AD, Li H, Eichler GS, Krausz KW, Weinstein JN, Fomace AJ, Gonzalez FJ, Idle JR (2008) UPLC-ESI-TOFMS-based metabolomics and gene expression dynamics inspector self-organizing metabolomic maps as tools for understanding the cellular response to ionizing radiation. Anal Chem 80:665–674PubMedCrossRefGoogle Scholar
  23. Pompella A, Visvikis A, Paolicchi A, De Tata V, Casini AF (2003) The changing faces of glutathione, a cellular protagonist. Biochem Pharm 66(8):1499–1503PubMedCrossRefGoogle Scholar
  24. Potapenko DI, Bagryanskaya EG, Grigoriev IA, Maksimov AM, Reznikov VA, Platonov VE, Clanton TL, Khramtsov VV (2005) Quantitative determination of SH groups using 19F NMR spectroscopy and disulfide of 2, 3, 5, 6-tetrafluoro-4-mercaptobenzoic acid. Magn Reson Chem 43:902–909PubMedCrossRefGoogle Scholar
  25. Rodriguez-Ariza A, Toribio F, Lopez-Barea J (1994) Rapid determination of glutathione status in fish liver using high-performance liquid chromatography and electrochemical detection. J Chrom B 656:311–318CrossRefGoogle Scholar
  26. Rover JL, Kubota LT, Hoehr NF (2001) Development of an amperometric biosensor based on glutathione peroxidase immobilized in a carbodiimide matrix for the analysis of reduced glutathione from serum. Clin Chim Acta 308:55–67PubMedCrossRefGoogle Scholar
  27. Sahlin E, Jagner D, Ratana-ohpas R (1997) Mercury nucleation on glassy carbon electrodes. Anal Chim Acta 346:157–164CrossRefGoogle Scholar
  28. Saitoh K, Yamada N, Ishikawa E, Nakajima H, Shibukawa M (2006) On-line redox derivatization liquid chromatography using double separation columns and one derivatization unit. J Sep Sci 29:49–56PubMedCrossRefGoogle Scholar
  29. Saitoh K, Koichi K, Yabiku F, Noda Y, Porter MD, Shibukawa M (2008) On-column electrochemical redox derivatization for enhancement of separation selectivity of liquid chromatography: use of redox reaction as secondary chemical equilibrium. J Chromatogr A 1180:66–72PubMedCrossRefGoogle Scholar
  30. Satoh T, Yoshioka Y (2006) Contribution of reduced and oxidized glutathione to signals detected by magnetic resonance spectroscopy as indicators of local brain redox state. Neurosci Res 55:34–39PubMedCrossRefGoogle Scholar
  31. Shertzer HG, Vasiliou V, Liu RM, Tabor MW, Nebert DW (1995) Enzyme induction by l-buthionine (SR)-sulfoximine in cultured mouse hepatoma cells. Chem Res Toxicol 8:431–436PubMedCrossRefGoogle Scholar
  32. Terpstra M, Marjanska M, Henry PG, Tkac I, Gruetter R (2006) Detection of an antioxidant profile in the human brain in vivo via double editing with MEGA-PRESS. Magn Reson Med 56:1192–1199PubMedCrossRefGoogle Scholar
  33. Valko M, Leibfritz D, Moncol J, Cronin MT, Mazur M, Telser J (2007) Free radicals and antioxidants in normal physiological functions and human disease. Int J Biochem Cell Biol 39:44–84PubMedCrossRefGoogle Scholar
  34. Wu HP (1994) Nature and stability of mercury thin films on glassy carbon electrodes under fast-scan anodic stripping voltammetry. Anal Chem 66(19):3151–3157CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  1. 1.Chemistry Department, Faculty of Arts and ScienceNamık Kemal UniversityTekirdağTurkey
  2. 2.Biochemistry Department, Faculty of ScienceEge UniversityIzmirTurkey

Personalised recommendations