Time Matters: the Toxicity of Zinc Oxide Nanoparticles to Lemna minor L. Increases with Exposure Time

  • Xiaolin Chen
  • John O’Halloran
  • Marcel A. K. Jansen


The use of zinc oxide nanoparticles (nano-ZnO) has rapidly increased in recent years, and this has triggered the need for versatile toxicity tests that can be used to test a range of different exposure scenarios. Acute exposure studies, using a variety of plant species, have overwhelmingly demonstrated nano-ZnO-induced toxicity, but substantial differences in the degree of phytotoxicity are reported in different studies. Here, we analysed the role of exposure time in determining the variation in phytotoxic effects. Using the model species Lemna minor, the effects of short-term (24 h), standardised (1 week) and chronic (up to 6 weeks) nano-ZnO exposure were compared. Nano-ZnO effects on Lemna minor growth indicators (biomass growth rate, root length), chlorophyll content and photosynthetic efficiency were measured. Rapid inhibitory effects of nano-ZnO on the maximal quantum yield of photosystem II could be measured after just 24-h exposure. Standardised (1 week) experiments revealed phytotoxic effects on Lemna minor biomass growth. More severe inhibitory effects on growth developed gradually over 4 to 6 weeks exposure to nano-ZnO, and these were qualitatively associated with increased zinc content in the plant. Such dynamics of nano-ZnO toxicity have not been elucidated before, and this study emphasises the importance of exposure time in studies of nanoparticle toxicity. We conclude that short-term, standardised experiments can potentially underestimate the environmental phytotoxicity, which may result from chronic exposure to nano-ZnO.


Lemna minor Nano-ZnO Chronic toxicity Acute toxicity 



X.C. gratefully acknowledges the support by the CSC (China Scholarship Council). M.A.K.J. acknowledges the support by W.o.B. The authors appreciate the technical assistance (Atomic Absorption Spectroscopy) from Dr. Qiushi Xie and help with the physico-chemical characterisation of particles from Dr. Guillaume Yuhel.


  1. Adhikari, T., Kundu, S., Biswas, A. K., Tarafdar, J. C., & Subba Rao, A. (2015). Characterization of zinc oxide nano particles and their effect on growth of maize (Zea mays L.) plant. Journal of Plant Nutrition, 38(10), 1505–1515.CrossRefGoogle Scholar
  2. Antoine, M., Florence, M., Éric, P., Gauthier, L., & Emmanuel, F. (2017). Environmental impact of engineered carbon nanoparticles: from releases to effects on the aquatic biota. Current Opinion in Biotechnology, 46(2017), 1–6.Google Scholar
  3. Ashby, E., Wangermann, E., & Winter, E. J. (1949). Studies in the morphogenesis of leaves. New Phytologist, 48(3), 374–381.CrossRefGoogle Scholar
  4. Boxall, A., Chaudhry, Q., Sinclair, C., Jones, A., Jefferson, B., & Watts, C. (2007). Current and future predicted environmental exposure to engineered nanoparticles. York: CSL.Google Scholar
  5. Brain, R. A., & Solomon, K. R. (2007). A protocol for conducting 7-day daily renewal tests with Lemna gibba. Nature Protocols, 2(4), 979–987.CrossRefGoogle Scholar
  6. Chang, Y. N., Zhang, M., Xia, L., Zhang, J., & Xing, G. (2012). The toxic effects and mechanisms of CuO and ZnO nanoparticles. Materials, 5(12), 2850–2871.CrossRefGoogle Scholar
  7. Chen, X., O’Halloran, J., & Jansen, M. A. K. (2016). The toxicity of zinc oxide nanoparticles to Lemna minor (L.) is predominantly caused by dissolved Zn. Aquatic Toxicology, 174(2016), 46–53.CrossRefGoogle Scholar
  8. Dastjerdi, R., & Montazer, M. (2010). A review on the application of inorganic nano-structured materials in the modification of textiles: focus on anti-microbial properties. Colloids and Surfaces. B, Biointerfaces, 79(1), 5–18.CrossRefGoogle Scholar
  9. Deng, X., Luan, Q., Chen, W., Wang, Y., Wu, M., Zhang, H., & Jiao, Z. (2009). Nanosized zinc oxide particles induce neural stem cell apoptosis. Nanotechnology, 20(11), 115101.CrossRefGoogle Scholar
  10. Dhawan, A., & Sharma, V. (2010). Toxicity assessment of nanomaterials: methods and challenges. Analytical and Bioanalytical Chemistry, 398(2), 589–605.CrossRefGoogle Scholar
  11. Dimkpa, C. O., Calder, A., Britt, D. W., McLean, J. E., & Anderson, A. J. (2011). Responses of a soil bacterium, Pseudomonas chlororaphis O6 to commercial metal oxide nanoparticles compared with responses to metal ions. Environmental Pollution, 159(7), 1749–1756.CrossRefGoogle Scholar
  12. 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(1), 36–43.CrossRefGoogle Scholar
  13. El Badawy, A. M., Luxton, T. P., Silva, R. G., Scheckel, K. G., Suidan, M. T., & Tolaymat, T. M. (2010). Impact of environmental conditions (pH, ionic strength, and electrolyte type) on the surface charge and aggregation of silver nanoparticles suspensions. Environmental Science and Technology, 44(4), 1260–1266.CrossRefGoogle Scholar
  14. Franklin, N., & Rogers, N. (2007). Comparative toxicity of nanoparticulate ZnO, bulk ZnO, and ZnCl2 to a freshwater microalga (Pseudokirchneriella subcapitata): the importance of particle solubility. Environmental Science and Technology, 41(24), 8484–8490.CrossRefGoogle Scholar
  15. García-Hevia, L., Valiente, R., Martín-Rodríguez, R., Renero-Lecuna, C., González, J., Rodríguez-Fernández, L., … Fanarraga, M. L. (2016). Nano-ZnO leads to tubulin macrotube assembly and actin bundling, triggering cytoskeletal catastrophe and cell necrosis. Nanoscale, 8(21), 10963–10973.CrossRefGoogle Scholar
  16. Ghodake, G., Seo, Y. D., & Lee, D. S. (2011). Hazardous phytotoxic nature of cobalt and zinc oxide nanoparticles assessed using Allium cepa. Hazardous Materials, 186(1), 952–955.CrossRefGoogle Scholar
  17. Gottschalk, F., Sonderer, T., Scholz, R. W., & Nowack, B. (2009). Modeled environmental concentrations of engineered nanomaterials (TiO2, ZnO, Ag, CNT, fullerenes) for different regions. Environmental Science and Technology, 43(24), 9216–9222.CrossRefGoogle Scholar
  18. Gubbins, E. J., Batty, L. C., & Lead, J. R. (2011). Phytotoxicity of silver nanoparticles to Lemna minor L. Environmental Pollution, 159(6), 1551–1559.CrossRefGoogle Scholar
  19. Hajra, A., & Mondal, N. K. (2017). Effects of ZnO and TiO2 nanoparticles on germination, biochemical and morphoanatomical attributes of Cicer arietinum L. Energy, Ecology and Environment, 2(4), 277–288.CrossRefGoogle Scholar
  20. Hall, S., Bradley, T., Moore, J. T., Kuykindall, T., & Minella, L. (2009). Acute and chronic toxicity of nano-scale TiO2 particles to freshwater fish, cladocerans, and green algae, and effects of organic and inorganic substrate on TiO2 toxicity. Nanotoxicology, 3(2), 91–97.CrossRefGoogle Scholar
  21. Hernandez-Viezcas, J. A., Castillo-Michel, H., Servin, A. D., Peralta-Videa, J. R., & Gardea-Torresdey, J. L. (2011). Spectroscopic verification of zinc absorption and distribution in the desert plant Prosopis juliflora-velutina (velvet mesquite) treated with ZnO nanoparticles. Chemical Engineering, 170(1–3), 346–352.CrossRefGoogle Scholar
  22. Hossell, J. C., & Baker, J. H. (1979). Estimation of the growth rates of epiphytic bacteria and Lemna minor in a river. Freshwater Biology, 9(4), 319–327.CrossRefGoogle Scholar
  23. Hu, C., Liu, X., Li, X., & Zhao, Y. (2014). Evaluation of growth and biochemical indicators of Salvinia natans exposed to zinc oxide nanoparticles and zinc accumulation in plants. Environmental Science and Pollution Research International, 21(1), 732–739.CrossRefGoogle Scholar
  24. Inskeep, W. P., & Bloom, P. R. (1985). Extinction coefficients of chlorophyll a and b in N,N-dimethylformamide and 80% acetone. Plant Physiology, 77(2), 483–485.CrossRefGoogle Scholar
  25. Juhel, G., Batisse, E., Hugues, Q., Daly, D., van Pelt, F. N. A. M., O’Halloran, J., & Jansen, M. A. K. (2011). Alumina nanoparticles enhance growth of Lemna minor. Aquatic Toxicology, 105(3–4), 328–336.CrossRefGoogle Scholar
  26. Keller, A. A., Mcferran, S., Lazareva, A., & Suh, S. (2013). Global life cycle releases of engineered nanomaterials. Nanoparticle Research, 15(6), 1692–1694.CrossRefGoogle Scholar
  27. Kim, S., Lee, S., & Lee, I. (2012). Alteration of phytotoxicity and oxidant stress potential by metal oxide nanoparticles in Cucumis sativus. Water, Air, and Soil Pollution, 223(5), 2799–2806.CrossRefGoogle Scholar
  28. Kumari, M., Khan, S. S., Pakrashi, S., Mukherjee, A., & Chandrasekaran, N. (2011). Cytogenetic and genotoxic effects of zinc oxide nanoparticles on root cells of Allium cepa. Hazardous Materials, 190(1–3), 613–621.CrossRefGoogle Scholar
  29. Lahive, E., O’Halloran, J., & Jansen, M. A. K. (2011). Differential sensitivity of four Lemnaceae species to zinc sulphate. Environmental and Experimental Botany, 71(1), 25–33.CrossRefGoogle Scholar
  30. Lee, C. W., Mahendra, S., Zodrow, K., Li, D., Tsai, Y. C., Braam, J., & Alvarez, P. J. J. (2010). Developmental phytotoxicity of metal oxide nanoparticles to Arabidopsis thaliana. Environmental Toxicology and Chemistry / SETAC, 29(3), 669–675.CrossRefGoogle Scholar
  31. Lee, S., Kim, S., Kim, S., & Lee, I. (2012). Assessment of phytotoxicity of ZnO NPs on a medicinal plant, Fagopyrum esculentum. Environmental Science and Pollution Research International, 20(2), 848–854.CrossRefGoogle Scholar
  32. Lewis, M. A. (1995). Use of freshwater plants for phytotoxicity testing: a review. Environmental Pollution, 87(3), 319–336.CrossRefGoogle Scholar
  33. Li, N., Georas, S., Alexis, N., Fritz, P., Xia, T., Williams, M. A., et al. (2016). A work group report on ultrafine particles (American Academy of Allergy, Asthma & Immunology): why ambient ultrafine and engineered nanoparticles should receive special attention for possible adverse health outcomes in human subjects. Journal of Allergy and Clinical Immunology, 138(2), 386–396.CrossRefGoogle Scholar
  34. Lin, D., & Xing, B. (2007). Phytotoxicity of nanoparticles: inhibition of seed germination and root growth. Environmental Pollution, 150(2), 243–250.CrossRefGoogle Scholar
  35. Lin, D., & Xing, B. (2008). Root uptake and phytotoxicity of ZnO nanoparticles. Environmental Science and Technology, 42(15), 5580–5585.CrossRefGoogle Scholar
  36. Mahajan, P., Dhoke, S. K., & Khanna, A. S. (2011). Effect of nano-ZnO particle suspension on growth of mung (Vigna radiata) and gram (Cicer arietinum) seedlings using plant agar method. Nanotechnology, 2011(9), 1–7.Google Scholar
  37. Ma, H., Wallis, L. K., Diamond, S., Li, S., Canas-Carrell, J., & Parra, A. (2014). Impact of solar UV radiation on toxicity of ZnO nanoparticles through photocatalytic reactive oxygen species (ROS) generation and photo-induced dissolution. Environmental Pollution, 193(2014), 165–172.CrossRefGoogle Scholar
  38. Nowack, B., & Bucheli, T. D. (2007). Occurrence, behavior and effects of nanoparticles in the environment. Environmental Pollution, 150(1), 5–22.CrossRefGoogle Scholar
  39. OECD (2002). OECD guidelines for the testing of chemicals: revised proposal for a new guideline 221. Lemna sp. growth inhibition test. OECD.Google Scholar
  40. Pandey, A. C., Sanjay, S. S., & Yadav, R. S. (2010). Application of ZnO nanoparticles in influencing the growth rate of Cicer arietinum. Experimental Nanoscience, 5(6), 488–497.CrossRefGoogle Scholar
  41. Pirson, A., & Göllner, E. (1953). Beobachtungen zur Entwicklungsphysiologie der Lemna minor L. Flora, 140, 485–498.Google Scholar
  42. Poynton, H. C., Lazorchak, J. M., Impellitteri, C. a, Smith, M. E., Rogers, K., Patra, M., … Vulpe, C. D. (2011). Differential gene expression in Daphnia magna suggests distinct modes of action and bioavailability for ZnO nanoparticles and Zn ions. Environmental Science & Technology, 45(2), 762–768.CrossRefGoogle Scholar
  43. Priyadarshini, B., Behera, S. S., Rath, P. P., Sahoo, T. R., & Parhi, P. K. (2017). Adsorption of xylenol orange dye on nano ZnO: kinetics, thermodynamics and isotherm study. AIP Conference Proceedings, 1832, 50043.CrossRefGoogle Scholar
  44. Read, D. S., Matzke, M., Gweon, H. S., Newbold, L. K., Heggelund, L., Ortiz, M. D., et al. (2016). Soil pH effects on the interactions between dissolved zinc, non-nano- and nano-ZnO with soil bacterial communities. Environmental Science & Pollution Research, 23(5), 4120–4128.CrossRefGoogle Scholar
  45. Ren, G., Hu, D., Cheng, E. W. C., Vargas-Reus, M. A., Reip, P., & Allaker, R. P. (2009). Characterisation of copper oxide nanoparticles for antimicrobial applications. Antimicrobial Agents, 33(6), 587–590.CrossRefGoogle Scholar
  46. Tang, Y., Li, S., Lu, Y., Li, Q., & Yu, S. (2014). The influence of humic acid on the toxicity of nano-ZnO and Zn2+ to the Anabaena sp. Environmental Toxicology, 30(8), 895–904.CrossRefGoogle Scholar
  47. Waalewijn-Kool, P. L., Diez Ortiz, M., van Straalen, N. M., & van Gestel, C. A. M. (2013). Sorption, dissolution and pH determine the long-term equilibration and toxicity of coated and uncoated ZnO nanoparticles in soil. Environmental Pollution, 178(1), 59–64.CrossRefGoogle Scholar
  48. Wang, H., Wick, R. L., & Xing, B. (2009). Toxicity of nanoparticulate and bulk ZnO, Al2O3 and TiO2 to the nematode Caenorhabditis elegans. Environmental Pollution, 157(4), 1171–1177.CrossRefGoogle Scholar

Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  • Xiaolin Chen
    • 1
  • John O’Halloran
    • 1
    • 2
  • Marcel A. K. Jansen
    • 1
    • 2
  1. 1.School of Biological, Earth and Environmental SciencesUniversity College CorkCorkIreland
  2. 2.Environmental Research InstituteUniversity College CorkCorkIreland

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