Antioxidant Defense System Parameters in Isolated Fish Hepatocytes Exposed to Bisphenol a — Effect of Vitamin C


In this study, isolated hepatocytes of pearl mullet (Alburnus tarichi) were exposed to bisphenol A (BPA) at concentrations of 25, 50, 100, and 200 µM for 24 h. Moreover, an in vitro antioxidant concentration of vitamin C (50 µM) was administrated to the culture medium along with the BPA exposures. Next, the antioxidant defense system parameters were analyzed. According to the results, the highest concentration of BPA (200 µM) proved to be severely toxic for the cells. The increased activities of superoxide dismutase (SOD) and glutathione-S-transferase (GST), the fuctuated activities of glutathione peroxidase (GPx), and the decreased content of reduced glutathione (GSH) were compared to the control group after the BPA exposures. Vitamin C co-administration was found to cause further increases in the SOD, GPx, and GST activities in some of the experimental groups and vitamin C could not restore the GSH content. Malondialdehyde (MDA) levels were observed to be unaffected in all exposure groups. These results show that BPA causes alterations in the antioxidant defenses of the isolated fsh hepatocytes. In addition, vitamin C co-administration along with BPA was found to be non-protective against BPA-induced oxidative stress, consequently, aggravated a negative BPA impact.


  1. 1.

    Almroth, B. C. (2008) Oxidative damage in fsh used is biomarkers in feld and laboratory studies. Department of Zoology/Zoophysiology, Chalmers Reproservice, Göteborg University, Sweden,; 74 pp.

    Google Scholar 

  2. 2.

    Belfroid, A., Van Velzen, M., Van der Horst, B., Vethaak, D. (2002) Occurrence of bisphenol A in surface water and uptake in fsh: evaluation of feld measurements. Chemosphere 49, 97–103.

    CAS  PubMed  Google Scholar 

  3. 3.

    Berry, M. N., Friend, D. S. (1969) High yield preparation of isolated rat liver parenchymal cells. J. Cell Biol. 43, 506–520.

    CAS  PubMed  PubMed Central  Google Scholar 

  4. 4.

    Beutler, E. (1984) Red cell metabolism. A manual of biochemical methods, 3rd edn. Grune and Startton, sNew York, pp. 105–106.

    Google Scholar 

  5. 5.

    Bindhumol, V., Chitra, K. C., Mathur, P. P. (2003) Bisphenol A induces reactive oxygen species generation in the liver of male rats. Toxicology 188, 117–124.

    CAS  PubMed  Google Scholar 

  6. 6.

    Bradford, M. M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248–254.

    CAS  Google Scholar 

  7. 7.

    Crain, D., Eriksen, M., Iguchi, T., Jobling, S., Laufer, H., LeBlanc, G. A., Guilette Jr., L. J. (2007) An ecological assessment of bisphenol-A: evidence from comparative biology. Reprod. Toxicol. 24, 225–239.

    CAS  PubMed  Google Scholar 

  8. 8.

    Çakal Arslan, Ö., Parlak, H. (2008) Effects of bisphenol A on the embryonic development of sea urchin (Paracentrotus lividus). Environ. Toxicol. 23, 387–392.

    Google Scholar 

  9. 9.

    Danulat, E., Selcuk, B. (1992) Life history and environmental conditions of the anadromous Chalcalburnus tarichi (Cyprinidae) in the highly alkaline Lake Van, Eastern Anatolia, Turkey. Arch. Hydrobiol. 126, 105–125.

    Google Scholar 

  10. 10.

    Das, K. K. Das, S. N., Ghundasi, S. A. (2008) Nickel, its adverse health effects and oxidative stress. Indian J. Med. Res. 128, 412–425.

    CAS  PubMed  Google Scholar 

  11. 11.

    Dickinson, D. A., Forman, H. J. (2002) Cellular glutathione and thiols metabolism. Biochem. Pharmacol. 64, 1019–1026.

    CAS  PubMed  Google Scholar 

  12. 12.

    Elp, M., Şen, F., Atıcı, A. A. (2014) The distribution area of tarek [Alburnus Tarichi (Guldenstaedtii, 1814)] in the Van lake basin, Turkey. Y.Y.U. J. Agr. Sci. 24, 228–232.

    Google Scholar 

  13. 13.

    El-Shenawy, N. S., Al-Ghamdi, O. A. (2014) Phenthoate induced-oxidative stress in fresh isolated mice hepatocytes: alleviation by ascorbic acid. Toxicol. Environ. Health Sci. 6, 67–80.

    Google Scholar 

  14. 14.

    Gangadharan, B., Murugan, M. A., Mathur, P. P. (2001) Effect of methoxychlor on antioxidant system of goat epididymal sperm in vitro. Asian J. Androl. 3, 285–288.

    CAS  PubMed  Google Scholar 

  15. 15.

    Habig, W. H., Pabst, M. J., Jakoby, W. B. (1974) Glutathione S-transferases. The frst enzymatic step in mercapturic acid formation. J. Biol. Chem. 249, 7130–7139.

    CAS  Google Scholar 

  16. 16.

    Honkanen, J. O., Holopainen, I. J., Kukkonen, J. V. K. (2004) Bisphenol A induces yolk-sac oedema and other adverse effects in landlocked salmon (Salmo salar m. sebago) yolk-sac fry. Chemosphere 55, 187–196.

    CAS  PubMed  Google Scholar 

  17. 17.

    Hulak, M., Gazo, I., Anna, S., Pavla, L. (2013) In vitro effects of bisphenol A on the quality parameters, oxidative stres, DNA integrity and adenosine triphosphate content in sterlet (Acipenser ruthe-nus) spermatozoa. Comp. Biochem. Physiol. 158, 64–71.

    CAS  Google Scholar 

  18. 18.

    Isik, I., Celik, I. (2008) Acute effects of methyl parathion and diazinon as inducers for oxidative stress on certain biomarkers in various tissues of rainbow trout (Oncorhynchus mykiss). Pest. Biochem. Physiol. 92, 38–42.

    CAS  Google Scholar 

  19. 19.

    Jain, S. K., McVie, R., Duett, J., Herbst, J. J. (1989) Erythrocyte membrane lipid peroxidation and glycolylated hemoglobin in diabetes. Diabetes 38, 1539–1543.

    CAS  PubMed  Google Scholar 

  20. 20.

    Jialal, I., Grundy, S. M. (1991) Preservation of the endogenous antioxidants in low density lipoprotein by ascorbate but not probucol during oxidative modifcation. J. Clin. Invest. 87, 43–53.

    Google Scholar 

  21. 21.

    Kabuto, H., Hasuike, S., Minagawa, N., Shishibori, T. (2003) Effects of bisphenol A on the metabolisms of active oxygen species in mouse tissues. Environ. Res. 93, 31–35.

    CAS  PubMed  Google Scholar 

  22. 22.

    Kaptaner, B. (2015) Cytotoxic effects of 4-octylphenol on fsh hepatocytes. Cytotechnology DOI: 10.1007/s10616-015-9916-3.

    Google Scholar 

  23. 23.

    Kaptaner, B., Kankaya, E. (2016) Caspase-3 activation in cytotoxicity of isolated rainbow trout (Oncorhycus mykiss) hepatocytes induced by bisphenol A. Fresen. Environ. Bull. 25, 1167–1174.

    CAS  Google Scholar 

  24. 24.

    Korkmaz, A., Kolankaya, D. (2009) The protective effect of ascorbic acid against renal ischemia-reperfusion injury in male rats. Ren. Fail. 31, 36–43.

    CAS  PubMed  Google Scholar 

  25. 25.

    Korkmaz, A., Ahbap, M. A., Kolankaya, D., Barlas, N. (2010) Infuence of vitamin C on bisphenol A, nonylphenol and octylphenol induced oxidative damages in liver of male rats. Food Chem. Toxicol. 48, 2868–2871.

    Google Scholar 

  26. 26.

    Kuch, H. M., Ballschmiter, K. (2001) Determination of endocrine-disrupting phenolic compounds and estrogens in surface and drinking water by HRGC-(NCI)-MS in the picogram per liter range. Environ. Sci. Technol. 35, 3201–3206.

    CAS  PubMed  Google Scholar 

  27. 27.

    Latchoumycandane, C., Chitra, K. C., Mathur, P. P. (2002) The Effect of 2,3,7,8,-tetrachlorodibenzo-p-dioxin on the antioxidant system in mitochondrial and microsomal fractions of rat testis. Toxicology 171, 113–118.

    Google Scholar 

  28. 28.

    Liu, C., Yu, K., Shi, X., Wang, J., Lam, P. K. S., Wu, R. S. S., Zhou, B. (2007) Induction of oxidative stress and apoptosis by PFOS and PFOA in primary cultured hepatocytes of freshwater tilapia (Oreochromis niloticus). Aquat. Toxicol. 82, 135–143.

    CAS  PubMed  Google Scholar 

  29. 29.

    Lv, X., Zhou, Q. F., Song, M. Y., Jiang, G. B., Shao, J. (2007) Vitellogenic reponses of 17 beta-estradiol and bisphenol A in male Chinese loach (Misgurnus anguillicaudatus). Environ. Toxicol. Pharmacol. 24, 155–159.

    CAS  PubMed  Google Scholar 

  30. 30.

    Mortensen, A. S., Tolfsen, C. C., Arukwe, A. (2006) Gene expression patterns in estrogen (nonylphenol) and aryl hydrocarbon receptor agonists (PCB-77) interaction using rainbow trout (Oncorhynchus mykiss) primary hepatocyte culture. J. Toxicol. Environ. Health 69, 1–19.

    Google Scholar 

  31. 31.

    Orta, B., Erkan, M. (2014) Effects of vitamin C on antioxidant systems and steroidogenic enzymes in sodium fuoride-exposed TM4 Sertoli cells. Fluoride 47, 139–151.

    CAS  Google Scholar 

  32. 32.

    Paglia, D. E., Valentine, W. N. (1967) Studies on the quantitative and qualitative and qualitative characterization of eryhrocyte glutathione peroxidase. J. Lab. Clin. Med. 70, 158–169.

    CAS  PubMed  PubMed Central  Google Scholar 

  33. 33.

    Park, C. B., Kim, B. H., Na, O. S., Choi, Y. C., Lee, Y. D., Baek, H. J., Kim, H. B., Takemura, A. (2007) Induction of in vitro vitellogenin synthesis by bisphenol, nonylphenol and octyphenol in Chinese minnow (Phoxinusoxy cephalus) hepatocytes. Korean J. Biol. Sci. 7, 227–235.

    Google Scholar 

  34. 34.

    Petersen, K., Tollefsen, K. (2011) Assessing combined toxicity of estrogen receptor agonists in a primary culture of rainbow trout (Oncorhynchus mykiss) hepatocytes. Aquat. Toxicol. 101, 186–195.

    CAS  PubMed  Google Scholar 

  35. 35.

    Rankhouhi, T. R., van Holsteijn, I., Letcher, R., Giesy, J. P., Van den Berg, M. (2002) Effects of primary exposure to environmental and natural estrogens on vitellogenin production in carp (Cyprinus carpio) hepatocytes. Toxicol. Sci. 67, 75–80.

    Google Scholar 

  36. 36.

    Rietjens, I., Boersma, M., de Haan, L., Spenkelink, B., Awad, H. M., Cnubben, N. H., van Zanden, J. J., van der Woude, H., Alink, G. M., Koeman, J. H. (2002) The pro-oxidant chemistry of the natural antioxidants vitamin C, vitamin E, carotenoids and favonoids. Environ. Toxicol. Pharmacol. 11, 321–333.

    CAS  PubMed  Google Scholar 

  37. 37.

    Sakagami. H., Satoh, K. (1997) Prooxidant action of two antioxidants: ascorbic acid and gallic acid. Anticancer Res. 17, 221–224.

    CAS  PubMed  Google Scholar 

  38. 38.

    Segner, H. (1198) Isolation and primary culture of teleost hepatocytes. Comp. Biochem. Physiol. 120, 71–81.

    Google Scholar 

  39. 39.

    Staples, C. A., Dorn, P. B., Klecka, G. M., O’Block, S. T., Harris, L. R. (1998) A review of the environmental fate, effects and exposures of bisphenol A. Chemosphere 36, 2149–2173.

    CAS  PubMed  Google Scholar 

  40. 40.

    Stephensen, E., Sturve, J., Forlin, L. (2002) Effects of redox cycling compounds on glutathione content and activity of glutathione-related enzymes in rainbow trout liver. Comp. Biochem. Physiol. C 144, 191–196.

    Google Scholar 

  41. 41.

    Turker, H., Takemura, A. (2011) Effects of environmental contaminants and natural substances on vitellogenesis in tilapia hepatocytes. Turk. J. Fish Aquat. Sci. 11, 539–545.

    Google Scholar 

  42. 42.

    Turkish Statistical Institute (TÜİK). (2013) Fishery Statistics. Ankara: publication number: 4349, ISSN 1013-6177, pp. 66.

    Google Scholar 

  43. 43.

    Unal, G., Marquez, E. C., Feld, M., Stavropoulos, P., Callard, I. P. (2014) Isolation of estrogen receptor subtypes and vitellogenin genes: expression in female Chalcalburnus tarichi. Comp. Biochem. Physiol. B 172–173, 67–73.

    PubMed  Google Scholar 

  44. 44.

    Ünal, G., Türkoğlu, V., Oğuz, A. R., Kaptaner, B. (2007) Gonadal histology and some biochemical characteristics of Chalcalburnus tarichi (Pallas, 1811) having abnormal gonads. Fish Physiol. Biochem. 33, 153–165.

    Google Scholar 

  45. 45.

    Wu, M., Xu, H., Shen, Y., Qiu, W., Yang, M. (2011) Oxidative stress in zebrafsh embryos induced by short-term exposure to bisphenol A, nonylphenol, and their mixture. Environ. Toxicol. Chem. 30, 2335–2341.

    CAS  PubMed  Google Scholar 

  46. 46.

    Wu, M., Xu, H., Yang, M., Xu, H. (2011) Effects of chronic bisphenol A exposure on hepatic anti-oxidant parameters in medaka (Oryzias latipes). Toxicol. Environ. Chem. 93, 270–278.

    CAS  Google Scholar 

  47. 47.

    Zhao, A. J., Liu, H. Q., Zhang, A. N., Wang, X. D., Zhang, H. Q. (2011) Effect of BDE-209 on glutathione system in Carrasius auratus. Environ. Toxicol. Pharmacol. 32, 35–39.

    CAS  PubMed  Google Scholar 

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Correspondence to Burak Kaptaner.

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Kaya, Ö., Kaptaner, B. Antioxidant Defense System Parameters in Isolated Fish Hepatocytes Exposed to Bisphenol a — Effect of Vitamin C. BIOLOGIA FUTURA 67, 225–235 (2016).

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  • Alburnus tarichi
  • bisphenol A
  • vitamin C
  • isolated fsh hepatocytes
  • antioxidant defenses