Fish Physiology and Biochemistry

, Volume 39, Issue 4, pp 957–966 | Cite as

Catalasic activity in fish liver: improvement of the UV to visible analytic method

  • Séverine Paris-Palacios
  • Laurence Delahaut
  • Alexis Carreras
  • Marielle Thomas
  • Sylvie Biagianti-Risbourg


Antioxidative defenses and more especially catalasic activity (CAT) are studied in a large range of scientific research thematics. In environmental sciences, the problematic of oxidative stress is of great interest as pollutants can induce perturbations of redox homeostasis. Consequently, changes in antioxidative defenses levels in fish tissues and particularly in liver are used as potential biomarkers of pollution. In most studies, the CAT was assayed by following during 5 min the consumption of H2O2 in cytosolic buffered extracts at 240 nm (UV-method). This study proposed a development of this method in the visible, using permanganate and a 525-nm detection, which was more accurate, sensitive, and rapid. Moreover, the hepatic CAT of six different fish species [a cyclidae (Nimbochromis linni), 3 cyprinidae (Brachydanio rerio, Rutilus rutilus, Cyprinus carpio), an anguillidae (Anguilla anguilla), and a percidae (Perca fluviatilus)] was evaluated with the two protocols (UV- and KMnO4-method). The results but also the thermal optimum of the reaction and the interest of CAT as biomarker in ecotoxicology were discussed.


Anguillidae Antioxidative defenses Catalase Cyclidae Cyprinidae Fish Liver Percidae Oxidative stress 



The present study was financially supported by the French National Research Agency through the program CIEL (ANR CESA005 2008).


  1. Ahmad I, Hamid T, Fatima M, Chand HS, Jain SK, Altran M, Raisuddin S (2000) Induction of hepatic antioxidants in freshwater catfish (Channa punctatus Bloch) is a biomarker of paper mill effluent exposure. Biochem Biophys Acta 15(23):37–48Google Scholar
  2. Ahmad I, Pacheco M, Santos MA (2004) Enzymatic and non enzymatic antioxidants as an adaptation to phagocyte-induced damage in Anguilla anguilla L. following in situ harbour water exposure. Ecotoxicol Environ Saf 57:290–302CrossRefPubMedGoogle Scholar
  3. Ahmad I, Oliveira M, Pacheco M, Santos MA (2005) Anguilla anguilla L. oxidative stress biomarkers responses to copper exposure with or without b-naphthoflavone pre-exposure. Chemosphere 61:267–275CrossRefPubMedGoogle Scholar
  4. Ahmad I, Pacheco M, Santos MA (2006) Anguilla anguilla L. oxidative stress biomarkers: an in situ study of freshwater wetland ecosystem (Pateira de Fermentelos, Portugal). Chemosphere 65(6):952–962CrossRefPubMedGoogle Scholar
  5. Almroth BC, Sturve J, Berglund A, Förlin L (2005) Oxidative damage in eelpout (Zoarces viviparous), measured as protein carbonyl and TBARS, as biomarkers. Aquat Toxicol 73:171–180CrossRefPubMedGoogle Scholar
  6. Aruoma OL (1998) Free radicals, oxidative stress and antioxidants in human health and disease. J Am Oil Chem 75:199–212CrossRefGoogle Scholar
  7. Babo S, Vasseur P (1992) In vitro effects of thiram on liver antioxidant enzyme activities of rainbow trout (Oncorhynchus mykiss). Aquat Toxicol 22(1):61–68CrossRefGoogle Scholar
  8. Beliaeff B, Burgeot T (2002) Integrated biomarker response: a useful tool for ecological risk assessment. Environ Toxicol Chem 21:1316–1322CrossRefPubMedGoogle Scholar
  9. Benedetti M, Fattorini D, Martuccio G, Nigro M, Regoli F (2009) Interactions between trace metals (Cu, Hg, Ni, Pb) and 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin in the Antarctic fish Trematomus bernacchii: oxidative effects on biotransformation pathway. Environ Toxicol Chem 28:818–825CrossRefPubMedGoogle Scholar
  10. Beutler E (1975) Red cell metabolism: a manual of biochemical methods. Grune and Straton, New YorkGoogle Scholar
  11. Bhatia S, Shukla R, Madhu SV, Gambhir JK, Prabhu KM (2003) Antioxidant status, lipid peroxidation and NO end products in patients of type 2 diabetes mellitus with nephropathy. Clin Biochem 36:557–562CrossRefPubMedGoogle Scholar
  12. Biagianti-Risbourg S, Paris-Palacios S, Mouneyrac C, Amiard-Triquet C (2012) Pollution acclimation, adaptation, resistance and tolerance in ecotoxicology. Encyclopedia of aquatic ecotoxicology In: Férard JF and Blaise C (eds) Springer, pp 135–146Google Scholar
  13. Bradford MM (1976) A rapid sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254CrossRefPubMedGoogle Scholar
  14. Broeg K, Westernhagen HV, Zander S, Körting W, Koehler A (2005) The “bioeffect assessment index”—a concept for the quantification of effects of marine pollution by an integrated biomarker approach. Mar Pollut Bull 50:495–503CrossRefPubMedGoogle Scholar
  15. Cavalheiro de Menezes C, Braga da Fonseca M, Pretto A, Dos Santos Miron D, Santi A (2011) Oxidative parameters of Rhamdia quelen in response to commercial herbicide containing clomazone and recovery pattern. Pestic Biochem Physiol 100(2):145–150CrossRefGoogle Scholar
  16. Celi P (2011) Biomarkers of oxidative stress in ruminant medicine. Immunopharm Immunotox 33(2):233–240CrossRefGoogle Scholar
  17. Cohen G, Dembiec D, Marcus J (1970) Measurement of catalase activity in tissue extracts. Anal Biochem 34:30–38CrossRefPubMedGoogle Scholar
  18. Crestani M, Menezes C, Glusczak L, Miron DS, Spanevello R, Silveira A, Gonçalves FF, Zanella R, Loro VL (2007) Effects of clomazone herbicide on biochemical and histological aspects of silver catfish (Rhamdia quelen) and recovery pattern. Chemosphere 67:2305–2311CrossRefPubMedGoogle Scholar
  19. Di Matteo V, Esposito E (2003) Biochemical and therapeutic effects of antioxidants in the treatment of Alzheimer’s disease, Parkinson’s disease, and amyotrophic lateral sclerosis. Curr Drug Targets CNS Neurol Disord 2:95–107CrossRefPubMedGoogle Scholar
  20. Galloway TS, Brown RJ, Browne MA, Dissanayake A, Lowe D, Jones MB, Depledge MH (2004) A multibiomarker approach to environmental assessment. Environ Sci Technol 38:1723–1731CrossRefPubMedGoogle Scholar
  21. Gerber M, Boutron-Ruault MC, Hercberg S, Riboli E, Scalbert A, Siess MH (2002) Food and cancer: state of the art about the protective effect of fruits and vegetables. Bull Cancer 89:293–312PubMedGoogle Scholar
  22. Gorbi S, Regoli F (2003) Total oxyradical scavenging capacity as an index of susceptibility to oxidative stress in marine organisms. Comments Toxicol 9:303–322CrossRefGoogle Scholar
  23. Góth L (1991) A simple method for determination of serum catalase activity and revision of reference range. Clin Chim Acta 196:143–152CrossRefPubMedGoogle Scholar
  24. Góth L, Németh H, Mészáros I (1984) Clinical study of the determination of serum catalase enzyme activity. Hung Sci Inst 57:7–12Google Scholar
  25. Johansson J, Hakan Borg LA (1988) A spectrophotometric method for determination of catalase activity in small tissue samples. Anal Biochem 174:331–336CrossRefPubMedGoogle Scholar
  26. Keramati V, Jamili S, Ramin M (2010) Effect of diazinon on catalase antioxidant enzyme activity in liver tissue of Rutilus rutilus. J Fish Aquat Sci 5(5):368–376CrossRefGoogle Scholar
  27. Krishnaiah D, Sarbatly R, Nithyanandam R (2011) A review of the antioxidant potential of medicinal plant species. Food Bioprod Proc 89(3):217–233CrossRefGoogle Scholar
  28. Lefer DJ, Granger DN (2000) Oxidative stress and cardiac disease. Am J Med 109:315–323CrossRefPubMedGoogle Scholar
  29. Leighton F, Poole B, Beaufay H, Baudhuin P, Coffey JW, Fowler S, De Duve C (1968) The large-scale separation of peroxisomes, mitochondria, and lysosomes from the livers of rats injected with Triton WR-1339. Improved isolation procedures, automated analysis, biochemical and morphological properties of fractions. J Cell Biol 37:482–513CrossRefPubMedGoogle Scholar
  30. Mao YX, Zheng H, Guo XQ, Chen R, Zheng WY (2002) Studies on the fluorometric method for determining activity of catalase and its application to marine biosamples. Chem J Chinese Univ 23(10):18–67Google Scholar
  31. Masuoka N, Wakimoto M, Ubuka T, Nakano T (1996) Spectrophotometric determination of hydrogen peroxide: catalase activity and rates of hydrogen peroxide removal by erythrocytes. Clin Chim Acta 254:101–112Google Scholar
  32. Modesto KA, Martinez CBR (2010) Roundup® causes oxidative stress in liver and inhibits acetylcholinesterase in muscle and brain of the fish Prochilodus lineatus. Chemosphere 78:294–299CrossRefPubMedGoogle Scholar
  33. Montavon P, Kukic KR, Bortlik K (2007) A simple method to measure effective catalase activities: optimization, validation, and application in green coffee. Anal Biochem 360(2):207–215CrossRefPubMedGoogle Scholar
  34. Moraes BS, Loro VL, Glusczak L, Pretto A, Menezes C, Marchezan E, Machado SO (2007) Effects of four rice herbicides on some metabolic and toxicology parameters of teleost fish (Leporinus obtusidens). Chemosphere 68:1597–1601CrossRefPubMedGoogle Scholar
  35. Mueller S, Riedel HD, Stremmel W (1997a) Determination of catalase activity at physiological hydrogen peroxide concentrations. Anal Biochem 245:55–60CrossRefPubMedGoogle Scholar
  36. Mueller S, Riedel HD, Stremmel W (1997b) Direct evidence for catalase as the predominant H2O2-removing enzyme in human erythrocytes. Blood 15:4973–4978Google Scholar
  37. Paris-Palacios S, Biagianti-Risbourg S, Vernet G (2000a) Biochemical and (ultra)structural hepatic perturbations of Brachydanio rerio (Teleostei, Cyprinidae) exposed to two sublethal concentrations of copper sulfate. Aquat Toxicol 50(1–2):109–124CrossRefPubMedGoogle Scholar
  38. Paris-Palacios S, Biagianti-Risbourg S, Fouley A, Vernet G (2000b) Metallothioneins in liver of Rutilus rutilus exposed to Cu2+. Analysis by metal summation, SH determination and spectrofluorimetry. Comp Biochem Physio C 126(2):113–122Google Scholar
  39. Paris-Palacios S, Biagianti-Risbourg S, Vernet G (2003) Metallothionein induction related to hepatic structural perturbations and antioxidative defences in roach (Rutilus rutilus) exposed to the fungicide procymidone. Biomarkers 8(2):128–141CrossRefPubMedGoogle Scholar
  40. Peuchant E, Brun J, Rigalleau V, Dubourg L, Thomas M, Daniel J (2004) Oxidative and antioxidative status in pregnant women with either gestational or type 1 diabetes. Clin Biochem 37:293–298CrossRefPubMedGoogle Scholar
  41. Piva F, Francesco Ciaprini F, Onorati F, Benedetti M, Fattorini D, Ausili A, Regoli F (2011) Assessing sediment hazard through a weight of evidence approach with bioindicator organisms: a practical model to elaborate data from sediment chemistry, bioavailability, biomarkers and ecotoxicological bioassays. Chemosphere 83(4):475–485CrossRefPubMedGoogle Scholar
  42. Regoli F, Pellegrini D, Winston GW, Gorbi S, Giuliani S, Virno-Lamberti C, Bompadre S (2002) Application of biomarkers for assessing the biological impact of dredged materials in the Mediterranean: the relationship between antioxidant responses and susceptibility to oxidative stress in the red mullet (Mullus barbatus). Mar Pollut Bull 44:912–922CrossRefPubMedGoogle Scholar
  43. Regoli F, Frenzilli G, Bocchetti R, Annarumma F, Scancelli V, Fattorini D, Nigro M (2004) Time-course variations of oxyradical metabolism, DNA integrity, and lysosomal stability in mussels, Mytilus galloprovincialis, during a field translocation experiment. Aquat Toxicol 68:167–178CrossRefPubMedGoogle Scholar
  44. Sayeed I, Parvez S, Pandey S, Bin-Hafeez B, Rizwanul H, Raisuddin S (2003) Oxidative stress biomarkers of exposure to deltamethrin in freshwater fish, Channa punctatus bloch. Ecotoxicol Environ Saf 56:295–301CrossRefPubMedGoogle Scholar
  45. Scandalious JG (1997) Molecular biology of the antioxidant defence genes encoding catalases and superoxide dismutases in maize. In: Hatzios KK (ed) Regulation of enzymatic systems detoxifying xenobiotics in plants. Kluver Academic, Dordrecht, pp 97–108CrossRefGoogle Scholar
  46. Shahidi F, Janitha PK, Wanasundara PD (1992) Phenolic antioxidants. Crit Rev Food Sci Nutr 32:67–103CrossRefPubMedGoogle Scholar
  47. Slaughter MR, O’Brien PJ (2000) Fully-automated spectrophotometric method for measurement of antioxidant activity of catalase. Clin Biochem 33(7):525–534CrossRefPubMedGoogle Scholar
  48. Smith MA, Rottkamp CA, Nunomura A, Raina AK, Perry G (2000) Oxidative stress in Alzheimer’s disease. Biochem Biophys Acta 1502:139–144CrossRefPubMedGoogle Scholar
  49. Sreejayan N, Rao M (1996) Free radical scavenging activity of Curcuminoids. Drug Res 46:169–171Google Scholar
  50. Steer P, Milligard J, Sarabi DM, Wessby B, Kahan T (2002) Cardiac and vascular structure and function are related to lipid peroxidation and metabolism. Lipids 37:231–236CrossRefPubMedGoogle Scholar
  51. Sullivan JB (1992) Cyrogenics, oxidizers, reducing agents, and explosives. In: Sullivan JB, Krieger GR (eds) Hazardous materials toxicology. Clin. Principles Environ health. Baltimore, Williams & Wilkins, pp 1192–1201Google Scholar
  52. Toni C, Menezes CC, Loro VL, Clasen BE, Cattaneo R, Santi A, Pretto A, Zanella R, Leitemperger J (2010) Oxidative stress biomarkers in Cyprinus carpio exposed to commercial herbicide bispyribac-sodium. J Appl Toxicol 30:590–595CrossRefPubMedGoogle Scholar
  53. Uchida K (2000) Role of reactive aldehyde in cardiovascular diseases. Free Radical Biol Med 28:1685–1696CrossRefGoogle Scholar
  54. Van der Oost R, Beyer J, Vermeulen NPE (2003) Fish bioaccumulation and biomarkers in environmental risk assessment: a review. Environ Toxicol Pharmacol 13:57–149CrossRefPubMedGoogle Scholar
  55. Van Lente F, Pepoy M (1990) Coupled-enzyme determination of catalase activity in erythrocytes. Clin Chem 36:1339–1343PubMedGoogle Scholar
  56. Whaley-Connell A, McCullough PA, Sowers JR (2011) The role of oxidative stress in the metabolic syndrome. Rev Cardiovasc Med 12(1):21–29PubMedGoogle Scholar
  57. Wheeler CR, Salzman JA, Elsayed NB, Omaye ST, Korte DW (1990) Automated assays for superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase activity. Anal Biochem 184:193–199CrossRefPubMedGoogle Scholar
  58. Winston GW, DiGiulio RT (1991) Prooxidant and antioxidant mechanisms in aquatic organisms. Aquat Toxicol 19(2):137–161CrossRefGoogle Scholar
  59. Yildirim NC, Benzer F, Danabas D (2011) Evaluation of environmental pollution at Munzur river of tunceli applying oxidative stress biomarkers in Capoeta trutta (Heckel, 1843). J Plant Sci 21(1):66–71Google Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2012

Authors and Affiliations

  • Séverine Paris-Palacios
    • 1
  • Laurence Delahaut
    • 1
  • Alexis Carreras
    • 1
  • Marielle Thomas
    • 2
  • Sylvie Biagianti-Risbourg
    • 1
  1. 1.Laboratoire d’Ecologie-Ecotoxicologie, Faculté des Sciences, EA4689 Interaction Animal-Environnement (IAE)Université de Reims Champagne-ArdenneReims Cedex 2France
  2. 2.Unité de Recherche Animal et Fonctionnalités des Produits Animaux (UR AFPA)Université Henri Poincaré de Nancy 1NancyFrance

Personalised recommendations