Advertisement

Analytical and Bioanalytical Chemistry

, Volume 405, Issue 8, pp 2635–2642 | Cite as

Ratios of biliary glutathione disulfide (GSSG) to glutathione (GSH): a potential index to screen drug-induced hepatic oxidative stress in rats and mice

  • Lei Cao
  • Daniel Waldon
  • Yohannes Teffera
  • John Roberts
  • Mary Wells
  • Meghan Langley
  • Zhiyang ZhaoEmail author
Original Paper

Abstract

Hepatotoxicity of drug candidates is one of the major concerns in drug screening in early drug discovery. Detection of hepatic oxidative stress can be an early indicator of hepatotoxicity and benefits drug selection. The glutathione (GSH) and glutathione disulfide (GSSG) pair, as one of the major intracellular redox regulating couples, plays an important role in protecting cells from oxidative stress that is caused by imbalance between prooxidants and antioxidants. The quantitative determination of the GSSG/GSH ratios and the concentrations of GSH and GSSG have been used to indicate oxidative stress in cells and tissues. In this study, we tested the possibility of using the biliary GSSG/GSH ratios as a biomarker to reflect hepatic oxidative stress and drug toxicity. Four compounds that are known to alter GSH and GSSG levels were tested in this study. Diquat (diquat dibromide monohydrate) and acetaminophen were administered to rats. Paraquat and tert-butyl hydroperoxide were administered to mice to induce changes of biliary GSH and GSSG. The biliary GSH and GSSG were quantified using calibration curves prepared with artificial bile to account for any bile matrix effect in the LC–MS analysis and to avoid the interference of endogenous GSH and GSSG. With four examples (in rats and mice) of drug-induced changes in the kinetics of the biliary GSSG/GSH ratios, this study showed the potential for developing an exposure response index based on biliary GSSG/GSH ratios for predicting hepatic oxidative stress.

Keywords

Biliary GSH/GSSG Oxidative stress Hepatotoxicity LC–MS Quantitation 

Notes

Acknowledgments

We would like to thank Dr. Adria Colletti for her in vivo expertise and Jerold S. Harmatz for statistical advice.

References

  1. 1.
    Horii I, Yamada H, Kikkawa R, Yamamoto T, Fukushima T, Tomizawa K (2008) In: Sahu SC (ed) Hepatotoxicity. Wiley, ChichesterGoogle Scholar
  2. 2.
    McMillian M, Nie A, Parker JB, Leone A, Kemmerer M, Bryant S, Herlich J, Yieh L, Bittner A, Liu X, Wan J, Johnson MD, Lord P (2005) Drug-induced oxidative stress in rat liver from a toxicogenomics perspective. Toxicol Appl Pharmacol 207:171CrossRefGoogle Scholar
  3. 3.
    Kim JM (2012) In: AV Lyubimov (Ed) Encyclopedia of drug metabolism and interactions. Wiley, New YorkGoogle Scholar
  4. 4.
    Meister A (1994) Glutathione-ascorbic acid antioxidant system in animals. J Biol Chem 269:9397Google Scholar
  5. 5.
    Bursell SE, King GL (2000) The potential use of glutathionyl hemoglobin as a clinical marker of oxidative stress. Clin Chem 46:145Google Scholar
  6. 6.
    Briviba K, Klotz L, Sies H (1999) In: Methods in enzymology. Academic, New YorkGoogle Scholar
  7. 7.
    Jain A, Martensson J, Stole E, Auld PA, Meister A (1991) Glutathione deficiency leads to mitochondrial damage in brain. Proc Natl Acad Sci USA 88:1913CrossRefGoogle Scholar
  8. 8.
    Meloni M, Nicolay JF (2003) Dynamic monitoring of glutathione redox status in UV-B irradiated reconstituted epidermis: effect of antioxidant activity on skin homeostasis. Toxicol In Vitro 17:609CrossRefGoogle Scholar
  9. 9.
    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 39:44CrossRefGoogle Scholar
  10. 10.
    Meister A, Anderson ME (1983) Glutathione. Annu Rev Biochem 52:711CrossRefGoogle Scholar
  11. 11.
    Smith CV, Jaeschke H (1989) Effect of acetaminophen on hepatic content and biliary efflux of glutathione disulfide in mice. Chem Biol Interact 70:241CrossRefGoogle Scholar
  12. 12.
    Jaeschke H (1990) Glutathione disulfide formation and oxidant stress during acetaminophen-induced hepatotoxicity in mice in vivo: the protective effect of allopurinol. J Pharmacol Exp Ther 255:935Google Scholar
  13. 13.
    Camera E, Picardo M (2002) Analytical methods to investigate glutathione and related compounds in biological and pathological processes. J Chromatogr B Analyt Technol Biomed Life Sci 781:181–206CrossRefGoogle Scholar
  14. 14.
    Pastore A, Federici G, Bertini E, Piemonte F (2003) Analysis of glutathione: implication in redox and detoxification. Clinica Chimica Acta 333:19CrossRefGoogle Scholar
  15. 15.
    Iwasaki Y, Saito Y, Nakano Y, Mochizuki K, Sakata O, Ito R, Saito K, Nakazawa H (2009) Chromatographic and mass spectrometric analysis of glutathione in biological samples. J Chromatogr B 877:3309CrossRefGoogle Scholar
  16. 16.
    Monostori P, Wittmann G, Karg E, Túri S (2009) Determination of glutathione and glutathione disulfide in biological samples: an in-depth review. J Chromatogr B 877:3331CrossRefGoogle Scholar
  17. 17.
    Eberle D, Clarke R, Kaplowitz N (1981) Rapid oxidation in vitro of endogenous and exogenous glutathione in bile of rats. J Biol Chem 256:2115Google Scholar
  18. 18.
    Guan X, Hoffman B, Dwivedi C, Matthees DP (2003) A simultaneous liquid chromatography/mass spectrometric assay of glutathione, cysteine, homocysteine and their disulfides in biological samples. J Pharm Biomed Anal 31:251CrossRefGoogle Scholar
  19. 19.
    Hammermeister DE, Serrano J, Schmieder P, Kuehl DW (2000) Characterization of dansylated glutathione, glutathione disulfide, cysteine and cystine by narrow bore liquid chromatography/electrospray ionization mass spectrometry. Rapid Commun Mass Spectrom 14:503CrossRefGoogle Scholar
  20. 20.
    Norris RL, Eaglesham GK, Shaw GR, Smith MJ, Chiswell RK, Seawright AA, Moore MR (2001) A sensitive and specific assay for glutathione with potential application to glutathione disulphide, using high-performance liquid chromatography-tandem mass spectrometry. J Chromatogr B: Biomed Sci Appl 762:17CrossRefGoogle Scholar
  21. 21.
    Zhang F, Bartels MJ, Geter DR, Jeong YC, Schisler MR, Wood AJ, Kan L, Gollapudi BB (2008) Quantitation of glutathione by liquid chromatography/positive electrospray ionization tandem mass spectrometry. Rapid Commun Mass Spectrom 22:3608CrossRefGoogle Scholar
  22. 22.
    Yilmaz O, Keser S, Tuzcu M, Guvenc M, Cetintas B, Irtegun S, Tastan H, Sahin K (2009) A practical HPLC method to measure reduced (GSH) and oxidized (GSSG) glutathione concentrations in animal tissues. J Anim Vet Adv 8:343Google Scholar
  23. 23.
    Madhu C, Gregus Z, Cheng CC, Klaassen CD (1992) Identification of the mixed disulfide of glutathione and cysteinylglycine in bile: dependence on gamma-glutamyl transferase and responsiveness to oxidative stress. J Pharmacol Exp Ther 262:896Google Scholar
  24. 24.
    Lauterburg BH, Smith CV, Hughes H, Mitchell JR (1984) Biliary excretion of glutathione and glutathione disulfide in the rat. Regulation and response to oxidative stress. J Clin Invest 73:124CrossRefGoogle Scholar
  25. 25.
    Vidt DG, Bredemeyer A, Sapirstein E, Sapirstein LA (1959) Effect of ether anesthesia on the cardiac output, blood pressure, and distribution of blood flow in the albino rat. Circ Res 7:759CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Lei Cao
    • 1
  • Daniel Waldon
    • 1
  • Yohannes Teffera
    • 1
  • John Roberts
    • 1
  • Mary Wells
    • 1
  • Meghan Langley
    • 1
  • Zhiyang Zhao
    • 1
    Email author
  1. 1.Pharmacokinetics and Drug MetabolismAmgen Inc.CambridgeUSA

Personalised recommendations