Biologia Plantarum

, Volume 53, Issue 1, pp 75–84 | Cite as

Waterlogging induced oxidative stress and antioxidant activity in pigeonpea genotypes

  • D. Kumutha
  • K. Ezhilmathi
  • R. K. Sairam
  • G. C. Srivastava
  • P. S. Deshmukh
  • R. C. Meena
Original Papers


The objective of this study was to examine the role of antioxidant enzymes in waterlogging tolerance of pigeonpea (Cajanus cajan L. Halls) genotypes ICP 301 (tolerant) and Pusa 207 (susceptible). Waterlogging resulted in visible yellowing and senescence of leaves, decrease in leaf area, dry matter, relative water content and chlorophyll content in leaves, and membrane stability index in roots and leaves. The decline in all parameters was greater in Pusa 207 than ICP 301. Oxidative stress in the form of superoxide radical, hydrogen peroxide and thiobarbituric acid reactive substances (TBARS) contents initially decreased, however at 4 and 6 d of waterlogging it increased over control plants, probably due to activation of DPI-sensitive NADPH-oxidase. Antioxidant enzymes such as superoxide dismutase, ascorbate peroxidase, glutathione reductase and catalase also increased under waterlogging. The comparatively greater antioxidant enzyme activities resulting in less oxidative stress in ICP 301 could be one of the factor determining its higher tolerance to flooding as compared to Pusa 207. This study is the first to conclusively prove that waterlogging induced increase in ROS is via NADPH oxidase.

Additional key words

anoxia ascorbate peroxidase Cajanus cajan catalase glutathione reductase hydrogen peroxide hypoxia oxidative stress superoxide radical superoxide dismutase 



ascorbate peroxidase






days after anthesis


diphenylene iodonium chloride


glutathione reductase


membrane stability index


reactive oxygen species


relative water content


superoxide dismutase


thiobarbituric acid reactive substances


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Aebi, H.: Catalase in vitro.-Methods Enzymol. 105: 121–126, 1984.PubMedCrossRefGoogle Scholar
  2. Agarwal, S., Sairam, R.K., Srivastava, G.C., Tyagi, A., Meena, R.C.: Role of ABA, salicylic acid, calcium and hydrogen peroxide on antioxidant enzymes induction in wheat seedlings.-Plant Sci. 169: 559–570, 2005.CrossRefGoogle Scholar
  3. Albrecht, G., Wiedenroth, E.M.: Protection against activated oxygen following re-aeration of hypoxically pre-treated wheat roots. The response of the glutathione system.-J. exp. Bot. 45: 449–455, 1994.CrossRefGoogle Scholar
  4. Arnon, D.I.: Copper enzymes in isolated chloroplasts. Polyphenol oxidase in Beta vulgaris.-Plant Physiol. 24: 1–15, 1949.PubMedCrossRefGoogle Scholar
  5. Beauchamp, C., Fridovich, I.: Superoxide dismutase. Improved assays and an assay applicable to acrylamide gels.-Anal. Biochem. 44: 276–287, 1971.PubMedCrossRefGoogle Scholar
  6. Biemelt, S., Keetman, U. Albrecht, G.: Re-aeration following hypoxia or anoxia leads to activation of the antioxidative defense system in roots of wheat seedlings.-Plant Physiol. 116: 651–658, 1998.PubMedCrossRefGoogle Scholar
  7. Biemelt, S., Keetman, U., Mock, H.P., Grimm, B.: Expression and activity of isoenzymes of superoxide dismutase in wheat roots in response to hypoxia and anoxia.-Plant Cell Environ. 23: 135–144, 2000.CrossRefGoogle Scholar
  8. Blokhina, O.B., Chirkova, T.V., Fagerstedt, K.V.: Anoxic stress leads to hydrogen peroxide formation in plant cells.-J. exp. Bot. 52: 1–12, 2001.CrossRefGoogle Scholar
  9. Blokhina, O.B., Fagerstedt, K.V., Chirkova, T.V.: Relationships between lipid peroxidation and anoxia tolerance in a range of species during post-anoxic reaeration.-Physiol. Plant. 105: 625–632, 1999.CrossRefGoogle Scholar
  10. Bowler, C., Montague, M.V., Inze, D.: Superoxide dismutase and stress tolerance.-Annu. Rev. Plant Physiol. Plant mol. Biol. 43: 83–116, 1992.CrossRefGoogle Scholar
  11. Chaitanya, K.S.K., Naithani, S.C.: Role of superoxide, lipid peroxidation and superoxide dismutase in membrane perturbation during loss of viability in seeds of Shorea robusta Gaertnf.-New Phytol. 126: 623–627, 1994.CrossRefGoogle Scholar
  12. Chirkova, T.V., Novitskaya, L.O., Blokhina, O.B.: Lipid peroxidation and antioxidant systems under anoxia in plants differing in their tolerance to oxygen deficiency.-Russ. J. Plant Physiol. 45: 55–62, 1998.Google Scholar
  13. Collaku, A., Harrison, S.A.: Loses in wheat due to waterlogging.-Crop Sci. 42: 444–450, 2002.CrossRefGoogle Scholar
  14. Crawford, R.M.M., Braendle, R.: Oxygen deprivation stress in a changing environment.-J. exp. Bot. 47: 145–159, 1996.CrossRefGoogle Scholar
  15. Crawford, R.M.M., Walton, J.C., Wollenweber-Ratzer, B.: Similarities between post-ischaemic injury to animal tissues and post anoxic injury in plants.-Proc. roy. Soc. Edinburgh 102B: 325–332, 1994.Google Scholar
  16. De Carvalho, M.C.C.G., Da Silva, D.C.G., Ruas, P.M., Medri, M.E., Ruas, E.A., Ruas, C.F.: Flooding tolerance and genetic diversity in populations of Luehea divaricata.-Biol. Plant. 52: 771–774, 2008.CrossRefGoogle Scholar
  17. Dhindsa, R.A., Plumb-Dhindsa, P., Thorpe, T.A.: Leaf senescence correlated with increased permeability and lipid peroxidation, and decreased levels of superoxide dismutase and catalase.-J. exp. Bot. 126: 93–101, 1981.CrossRefGoogle Scholar
  18. Drew, MC.: Oxygen deficiency and root metabolism: injury and acclimation under hypoxia and anoxia.-Annu. Rev. Plant Physiol. Plant mol. Biol. 48: 223–250, 1997.PubMedCrossRefGoogle Scholar
  19. Else, M.A., Davies, W.S., Malone, M., Jackson, M.S.: A negative hydraulic message from oxygen-deficient roots of tomato plant?-Plant Physiol. 109: 1017–1024, 1995.PubMedGoogle Scholar
  20. Elstner, E.F.: Metabolism of activated oxygen species.-In: Davies, D.D. (ed.): The Biochemistry of Plants. Biochemistry of Metabolism. Vol. 11. Pp. 253–315. Academic Press, San Diego 1986.Google Scholar
  21. Fukao, T., Bailey-Serres, J.: Plant responses to hypoxia is survival a balancing act.-Trends Plant Sci. 9: 449–456, 2004.PubMedCrossRefGoogle Scholar
  22. Gambrell, R.P., Patrick, W.H.: Chemical and microbiological properties of anaerobic soils and sediments.-In: Hook, D.D., Crawford, R.M.M. (ed.): Plant Life in Anaerobic Environments. Pp. 375–423. Ann Arbor Scientific Publications, Ann Arbor 1978.Google Scholar
  23. Heath, R.L., Packer, L.: Photoperoxidation in isolated chloroplast. I. Kinetics and stoichiometry of fatty acid peroxidation.-Arch. Biochem. Biophys. 125: 189–198, 1968.PubMedCrossRefGoogle Scholar
  24. Hiscox, J.D., Israelstam, G.F.: A method for extraction of chloroplast from leaf tissue without maceration.-Can. J. Bot. 57: 1332–1334, 1979.CrossRefGoogle Scholar
  25. Jackson, M.B., Herman, B. Goodenogh, A.: An examination of the importance of ethanol in causing injury to flooded plants.-Plant Cell Environ. 5: 163–172, 1982.Google Scholar
  26. Jackson, M.B., Drew, M.C.: Effects of flooding on growth and metabolism of herbaceous plants.-In: Kozlowski, T.T. (ed.): Flooding and Plant Growth. Pp. 47–128. Academic Press, Orlando 1984.Google Scholar
  27. Kalashnikov, Yu.E., Balakhnina, T.I., Zakrzhevsky, D.A.: Effect of soil hypoxia on activation of oxygen and the system of protection from oxidative destruction in roots and leaves of Hordeum vulgare.-Russ. J. Plant Physiol. 41: 583–588, 1994.Google Scholar
  28. Kramer, P.J., Jackson, W.T.: Causes of injury to flooded tobacco plants.-Plant Physiol. 29: 241–245, 1954.PubMedCrossRefGoogle Scholar
  29. Laemmli, U.K.: Cleavage of structural proteins during the assembly of the head of bacteriophage T4.-Nature 227: 680–685, 1970.PubMedCrossRefGoogle Scholar
  30. Min, X.J., Bartholomew, D.P.: Effects of flooding and drought on ethylene metabolism, titratable acidity and fruiting of pineapple.-Acta Hort. 666: 135–148, 2005.Google Scholar
  31. Monk, L.S., Fagerstedt, K.V., Crawford, R.M.M.: Superoxide dismutase as an anaerobic polypeptide-a key factor in recovery from oxygen deprivation in Iris pseudacorus?-Plant Physiol. 85: 1016–1020, 1987.PubMedCrossRefGoogle Scholar
  32. Monk, L.S., Fagerstedt, K.V., Crawford, R.M.M.: Oxygen toxicity and superoxide dismutase as an antioxidant in physiological stress.-Physiol Plant. 76: 456–459, 1989.Google Scholar
  33. Naidoo, G.: Effects of flooding on leaf water potential and stomatal resistance in Bruguiera gymporrhiza (L.) Lam.-New Phytol. 93: 369–376, 1983.CrossRefGoogle Scholar
  34. Nakano, Y., Asada, K.: Hydrogen peroxide is scavenged by ascorbate specific peroxidase in spinach chloroplasts.-Plant Cell Physiol. 22: 867–880, 1981.Google Scholar
  35. Oberson, J., Pavelic, D., Braendle, R., Rawler, A.: Nitrate increases membrane stability of potato cells under anoxia.-J. Plant Physiol. 155: 792–794, 1999.Google Scholar
  36. Rao, M.V., Paliyath, G., Ormrod, D.P. Murr, D.P., Watkins, C.B.: Influence of salicylic acid on H2O2 production, oxidative stress and H2O2 metabolizing enzymes.-Plant Physiol. 115: 137–149, 1997.PubMedCrossRefGoogle Scholar
  37. Rawyler, A., Arpagaus, S., Braendle, R.: Impact of oxygen stress and energy availability on membrance stability of plant cells.-Ann. Bot. 90: 499–507, 2002.PubMedCrossRefGoogle Scholar
  38. Richard, B., Couce, I., Raymond, P., Saglio, P.H., Saint-Ges, V., Pradet, A.: Plant metabolism under hypoxia and anoxia.-Plant Physiol. Biochem. 32: 1–10, 1994.Google Scholar
  39. Sairam, R.K.: Effect of moisture stress on physiological activities of two contrasting wheat genotypes.-Indian J. exp. Biol. 32: 594–593, 1994.Google Scholar
  40. Sairam, R.K., Kumutha, D., Ezhilmathi, K., Deshmukh, P.S., Srivastava, G.C.: Physiology and biochemistry of waterlogging tolerance in plants.-Biol. Plant. 52: 401–412, 2008.CrossRefGoogle Scholar
  41. Sairam, R.K., Srivastava, G.C.: Water stress tolerance of wheat (Triticum aestivum L.): Variations in hydrogen peroxide accumulation and antioxidant activity in tolerant and susceptible genotypes.-J. Agron. Crop Sci. 186: 63–70, 2001.CrossRefGoogle Scholar
  42. Sairam, R.K., Rao, K.V., Srivastava, G.C.: Differential response of wheat genotypes to long term salinity stress in relation to oxidative stress, antioxidant activity and osmolyte concentration.-Plant Sci. 163: 1037–1046, 2002.CrossRefGoogle Scholar
  43. Sairam, R.K., Srivastava, G.C., Saxena, D.C.: Increased antioxidant activity under elevated temperatures: a mechanism of heat stress tolerance in wheat genotypes.-Biol. Plant. 43: 245–251, 2000.CrossRefGoogle Scholar
  44. Sandalio, L.M., Palma, P.M., Del-Rio, L.A.: Localization of manganese superoxide dismutase in peroxisomes isolated from Pisum sativum L.-Plant Sci. 51: 1–8, 1987.CrossRefGoogle Scholar
  45. Singh, K., Sharma, S.P., Singh, T.K., Singh, Y.: Effect of waterlogging on growth, yield and nutrient concentration of black gram and green gram under subtropical condition of Varanasi.-Ann. agr. Res. 7: 169–177, 1986.Google Scholar
  46. Smith, I.K., Vierheller, T.L., Thorne, C.A.: Assay of glutathione reductase in crude tissue homogenates using 5, 5′-dithiobis (2-nitrobenzoic acid).-Anal. Biochem. 175: 408–413, 1988.PubMedCrossRefGoogle Scholar
  47. Sorte, N.V., Deotah, R.D., Meshram, J.H., Chanekar, M.A.: Tolerance of soybean cultivars of waterlogging at various growth states.-J. Soil Crops 6: 68–72, 1996.Google Scholar
  48. Tadege, M., Dupuis, I., Kuhlemeier, C.: Ethanolic fermentation: new functions for an old pathway.-Trends Plant Sci. 4: 320–325, 1999.PubMedCrossRefGoogle Scholar
  49. Ushimaru, T., Maki, Y., Sano, S., Koshiba, K., Asada, K., Tsuji, H.: Induction of enzymes involved in the ascorbate-dependent antioxidative system, namely ascorbate peroxidase, mono dehydroascorbate reductase and dehydroascorbate reductase, after exposure to air of rice (Oryza sativa) seedlings germinated under water.-Plant Cell Physiol. 38: 541–549, 1997.Google Scholar
  50. Van Toai, T.T., Bolles, C.S.: Postanoxic injury in soybean (Glycine max) seedlings.-Plant Physiol. 97: 588–592, 1991.CrossRefGoogle Scholar
  51. Vartapetian, B.B., Jackson, M.B.: Plant adaptations to anaerobic stress.-Ann. Bot. 79(Suppl. A): 3–20, 1997.Google Scholar
  52. Weatherley, P.E.: Studies in the water relations of cotton plants. I. The field measurement of water deficit in leaves.-New Phytol. 49: 81–97, 1950.CrossRefGoogle Scholar
  53. Yan, B., Dai, Q., Liu, X., Huang, S., Wang, Z.: Flooding-induced membrane damage, lipid oxidation and activated oxygen generation in corn leaves.-Plant Soil 179: 261–268, 1996.CrossRefGoogle Scholar
  54. Yu, Q., Rengel, Z.: Drought and salinity differentially influence activities of superoxide dismutase in narrow-leafed lupines.-Plant Sci. 142: 1–11, 1999.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2009

Authors and Affiliations

  • D. Kumutha
    • 1
  • K. Ezhilmathi
    • 1
  • R. K. Sairam
    • 1
  • G. C. Srivastava
    • 1
  • P. S. Deshmukh
    • 1
  • R. C. Meena
    • 1
  1. 1.Indian Agricultural Research InstituteDivision of Plant PhysiologyNew DelhiIndia

Personalised recommendations