, Volume 20, Issue 4, pp 815–826 | Cite as

Biochemical responses of Lemna minor experimentally exposed to cadmium and zinc

  • Biljana Balen
  • Mirta Tkalec
  • Sandra Šikić
  • Sonja Tolić
  • Petra Cvjetko
  • Mirjana Pavlica
  • Željka Vidaković-Cifrek


The effects of 5 μM cadmium (Cd), a non-essential toxic element and 25 and 50 μM zinc (Zn), an essential micronutrient, were investigated in aquatic plant Lemna minor L. after 4 and 7 days of exposure to each metal alone or to their combinations. Both metals showed tendency to accumulate with time, but when present in combination, they reduced uptake of each other. Cd treatment increased the lipid peroxidation and protein oxidation indicating appearance of oxidative stress. However, Zn supplementation in either concentration reduced values of both parameters, while exposure to Zn alone resulted in elevated level of lipid peroxidation and protein oxidation but only on the 7th day. Enhanced DNA damage, which was found on the 4th day in plants treated with Cd alone or in combination with Zn, was reduced on the 7th day in combined treatments. Higher catalase activity obtained in all treated plants on the 4th day of experiment was reduced in Zn-treated plants, but remained high in plants exposed to Cd alone or in combination with Zn after 7 days. Cd exposure resulted in higher peroxidase activity, while Zn addition prominently reduced peroxidase activity in the plants subjected to Cd stress. In conclusion, Cd induced more pronounced oxidative stress and DNA damage than Zn in applied concentrations. Combined treatments showed lower values of oxidative stress parameters—lipid peroxidation, protein oxidation and peroxidase activity as well as lower DNA damage, which indicates alleviating effect of Zn on oxidative stress in Cd-treated plants.


Cadmium Zinc Lemna minor L. Oxidative stress DNA damage 



This work was supported by the Ministry of Science, Education and Sports of the Republic of Croatia, projects no. 119-1191196-1202 and 119-0982934-3110.


  1. Aebi M (1984) Catalase in vitro. Method Enzymol 105:121–126CrossRefGoogle Scholar
  2. Aravind P, Prasad MNV (2003) Zinc alleviates cadmium induced toxicity in Ceratophyllum demersum, a fresh water macrophyte. Plant Physiol Biochem 41:391–397CrossRefGoogle Scholar
  3. Aravind P, Prasad MNV (2005) Cadmium-zinc interactions in a hydroponic system using Ceratophyllum demersum L.: adaptive ecophysiology, biochemistry and molecular toxicology. Braz J Plant Physiol 17:3–20CrossRefGoogle Scholar
  4. Aravind P, Prasad MNV, Malec P, Waloszek A, Strzałka K (2009) Zinc protects Ceratophyllum demersum L. (free-floating hydrophyte) against reactive oxygen species induced by cadmium. J Trace Elem Med Biol 23:50–60CrossRefGoogle Scholar
  5. Arora A, Sairam RK, Srivastava GC (2002) Oxidative stress and antioxidative system in plants. Curr Sci 82:1227–1234Google Scholar
  6. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254CrossRefGoogle Scholar
  7. Bueno P, Piqueras A (2002) Effect of transition metals on stress, lipid peroxidation and antioxidant enzyme activities in tobacco cell cultures. Plant Growth Regul 36:161–167CrossRefGoogle Scholar
  8. Cakmak I (2000) Possible roles of zinc in protecting plant cells from damage by reactive oxygen species. New Phytol 146:185–205CrossRefGoogle Scholar
  9. Chance B, Maehly AC (1955) Assay of catalases and peroxidases. Method Enzymol 2:764–775CrossRefGoogle Scholar
  10. Chaoui A, Mazhoudi S, Ghorbal MH, El Ferjani E (1997) Cadmium and zinc induction of lipid peroxidation and effects on antioxidant enzyme activities in bean (Phaseolus vulgaris L.). Plant Sci 127:139–147CrossRefGoogle Scholar
  11. Cho U-H, Seo N-H (2005) Oxidative stress in Arabidopsis thaliana exposed to cadmium is due to hydrogen peroxide accumulation. Plant Sci 168:113–120CrossRefGoogle Scholar
  12. Cuypers A, Vangronsveld J, Clijsters H (2001) The redox status of plant cells (AsA and GSH) is sensitive to zinc imposed oxidative stress in roots and primary leaves of Phaseolus vulgaris. Plant Physiol Biochem 39:657–664CrossRefGoogle Scholar
  13. Das P, Samantaray S, Rout GR (1997) Studies on cadmium toxicity in plants: a review. Environ Pollut 98:29–36CrossRefGoogle Scholar
  14. Drost W, Matzke M, Backhaus T (2007) Heavy metal toxicity to Lemna minor: studies on the time dependence of growth inhibition and the recovery after exposure. Chemosphere 67:36–43CrossRefGoogle Scholar
  15. Gichner T, Patková Z, Száková J, Demnerová K (2004) Cadmium induces DNA damage in tobacco roots, but no DNA damage, somatic mutations or homologous recombination in tobacco leaves. Mutat Res 559:49–57Google Scholar
  16. Gichner T, Patková Z, Száková J, Žnidar I, Mukherjee A (2008) DNA damage in potato plants induced by cadmium, ethyl methanesulphonate and γ-rays. Environ Exp Bot 62:113–119CrossRefGoogle Scholar
  17. Gratão PL, Monteiro CC, Antunes AM, Peres LEP, Azevedo RA (2008) Acquired tolerance of tomato (Lycopersicon esculentum cv. Micro-Tom) plants to cadmium-induced stress. Ann Appl Biol 153:321–333CrossRefGoogle Scholar
  18. Hassan MJ, Zhang G, Wu F, Wie K, Chen Z (2005) Zinc alleviates growth inhibition and oxidative stress caused by cadmium in rice. J Plant Nutr Soil Sci 168:255–261CrossRefGoogle Scholar
  19. Heath RL, Packer L (1968) Photoperoxidation in isolated chloroplasts. I. Kinetics and stoichiometry of fatty acid peroxidation. Arch Biochem Biophys 125:189–198CrossRefGoogle Scholar
  20. Hiraga S, Sasaki K, Ito H, Ohashi Y, Matsui H (2001) A large family of class III plant peroxidases. Plant Cell Physiol 42:462–468CrossRefGoogle Scholar
  21. Horvat T, Vidaković-Cifrek Ž, Oreščanin V, Tkalec M, Pevalek-Kozlina B (2007) Toxicity assessment of heavy metal mixtures by Lemna minor L. Sci Total Environ 384:229–238CrossRefGoogle Scholar
  22. Hossain Z, Huq F (2002) Studies on the interaction between Cd2+ ions and DNA. J Inorg Biochem 90:85–96CrossRefGoogle Scholar
  23. ISO - International Organization for Standardization (2006) Determination of the toxic effect of water constituents and wastewater on duckweed (Lemna minor)—Duckweed growth inhibition test. ISO norm 20079Google Scholar
  24. Kawano T (2003) Role of the reactive oxygen species generating peroxidase reactions in plant defence and growth induction. Plant Cell Rep 21:829–837Google Scholar
  25. Khellaf N, Zerdaoui M (2010) Growth response of the duckweed Lemna gibba L. to copper and nickel phytoaccumulation. Ecotoxicology 19:1363–1368CrossRefGoogle Scholar
  26. Kwan KHM, Smith S (1991) Some aspects of the kinetics of cadmium uptake by fronds of Lemna minor L. New Phytol 117:91–102CrossRefGoogle Scholar
  27. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685CrossRefGoogle Scholar
  28. Levine RL, Williams JA, Stadtman ER, Shacter E (1994) Carbonyl assay for determination of oxidatively modified proteins. Method Enzymol 233:346–357CrossRefGoogle Scholar
  29. Lewis MA (1995) Use of freshwater plants for phytotoxicity testing: A review. Environ Pollut 87:319–336CrossRefGoogle Scholar
  30. Lin CW, Chang HB, Huang HJ (2005) Zinc induces mitogen-activated protein kinase activation by reactive oxygen species in rice roots. Plant Physiol Biochem 43:963–968CrossRefGoogle Scholar
  31. Lin A, Zhang X, Chen M, Cao Q (2007) Oxidative stress and DNA damages induced by cadmium accumulation. J Environ Sci 19:596–602CrossRefGoogle Scholar
  32. Mithöfer A, Schultze B, Boland W (2004) Biotic and heavy metal stress response in plants: evidence for common signals. FEBS Lett 566:1–5CrossRefGoogle Scholar
  33. Mittler R (2002) Oxidative stress, antioxidants and stress tolerance. Trends Plant Sci 7:405–410CrossRefGoogle Scholar
  34. Mohan BS, Hosetti BB (1997) Potential phytotoxicity of lead and cadmium to Lemna minor grown in sewage stabilization ponds. Environ Pollut 98:233–238CrossRefGoogle Scholar
  35. Muschitz A, Faugeron C, Morvan H (2009) Response of cultured tomato cells subjected to excess zinc: role of cell wall in zinc compartmentation. Acta Physiol Plant 31:1197–1204CrossRefGoogle Scholar
  36. Peraza MA, Fierro FA, Barber DS, Casarez E, Rael LT (1998) Effects of micronutrients on metal toxicity. Environ Health Persp 106:203–216CrossRefGoogle Scholar
  37. Pirson A, Seidel F (1950) Zell- und stoffwechselphysiologische Untersuchungen an der Wurzel von Lemna minor unter besonderer Berücksichtigung von Kalium- und Calciummangel [Cell metabolism and physiology in Lemna minor root deprived of potassium and calcium, in German]. Planta 38:431–473CrossRefGoogle Scholar
  38. Razinger J, Dermastia M, Dolenc Koce J, Zrimec A (2008) Oxidative stress in duckweed (Lemna minor L.) caused by short-term cadmium exposure. Environ Pollut 153:687–694CrossRefGoogle Scholar
  39. Rellán-Álvarez R, Ortega-Villasante C, Álvarez-Fernández A, Del Campo FF, Hernández LE (2006) Stress responses of Zea mays to cadmium and mercury. Plant Soil 279:41–50CrossRefGoogle Scholar
  40. Rout GR, Das P (2003) Effect of metal toxicity on plant growth and metabolism: I. Zinc. Agronomie 23:3–11CrossRefGoogle Scholar
  41. Sandalio LM, Dalurzo HC, Gómez M, Romero-Puertas MC, del Río LA (2001) Cadmium-induced changes in the growth and oxidative metabolism of pea plants. J Exp Bot 52:2115–2126Google Scholar
  42. Shah K, Kumar RG, Verma S, Dubey RS (2001) Effect of cadmium on lipid peroxidation, superoxide anion generation and activities of antioxidant enzymes in growing rice seedlings. Plant Sci 161:1135–1144CrossRefGoogle Scholar
  43. Shaw BP, Sahu SK, Mishra RK (2004) Heavy metal induced oxidative damage in terrestrial plants. In: Prasad MNV (ed) Heavy metal stress in plants. From biomolecules to ecosystems. Springer, Berlin, Heidelberg, pp 84–126Google Scholar
  44. Singh S, Eapen S, D’Souza SF (2006) Cadmium accumulation and its influence on lipid peroxidation and antioxidative system in an aquatic plant, Bacopa monnieri L. Chemosphere 62:233–246CrossRefGoogle Scholar
  45. Tkalec M, Prebeg T, Roje V, Pevalek-Kozlina B, Ljubešić N (2008) Cadmium-induced responses in duckweed Lemna minor L. Acta Physiol Plant 30:881–890CrossRefGoogle Scholar
  46. Tuteja N, Singh MB, Misra MK, Bhalla PL, Tuteja R (2001) Molecular mechanisms of DNA damage and repair: progress in plants. A review. Crit Rev Biochem Mol Biol 36:337–397CrossRefGoogle Scholar
  47. Ünyayar S, Celik A, Cekic OF, Gozel A (2006) Cadmium induced genotoxicity, cytotoxicity and lipid peroxidation in Allium sativum and Vicia faba. Mutagenesis 21:77–81CrossRefGoogle Scholar
  48. Vaillant N, Monnet F, Hitmi A, Sallanon H, Coudret A (2005) Comparative study of responses in four Datura species to a zinc stress. Chemosphere 59:1005–1013CrossRefGoogle Scholar
  49. Vallee BL, Falchuk KH (1993) The biochemical basis of zinc physiology. Physiol Rev 73:79–118Google Scholar
  50. Valverde M, Trejo C, Rojas E (2001) Is the capacity of lead acetate and cadmium chloride to induce genotoxic damage due to direct DNA-metal interaction? Mutagenesis 16:265–270CrossRefGoogle Scholar
  51. Welch RM (1995) Micronutrient nutrition of plants. Crit Rev Plant Sci 14:49–82Google Scholar
  52. Woodbury WA, Spencer K, Stahlmann MA (1971) An improved procedure using ferricyanide for detecting catalase isozymes. Anal Biochem 44:301–305CrossRefGoogle Scholar
  53. Wu F, Zhang G (2002) Alleviation of cadmium-toxicity by application of zinc and ascorbic acid in barley. J Plant Nutr 25:2745–2761CrossRefGoogle Scholar
  54. Źróbek-Sokolnik A, Asard H, Górska-Koplińska K, Górecki RJ (2009) Cadmium and zinc-mediated oxidative burst in tobacco BY-2 cell suspension cultures. Acta Physiol Plant 31:43–49CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Biljana Balen
    • 1
  • Mirta Tkalec
    • 2
  • Sandra Šikić
    • 3
  • Sonja Tolić
    • 3
  • Petra Cvjetko
    • 1
  • Mirjana Pavlica
    • 1
  • Željka Vidaković-Cifrek
    • 2
  1. 1.Department of Molecular Biology, Faculty of ScienceUniversity of ZagrebZagrebCroatia
  2. 2.Department of Botany and Botanical Garden, Faculty of ScienceUniversity of ZagrebZagrebCroatia
  3. 3.Department of EcologyInstitute of Public Health “Dr. Andrija Štampar”ZagrebCroatia

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