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Superoxide production by thylakoids during chilling and its implication in the susceptibility of plants to chilling-induced photoinhibition

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Factors influencing the rate of superoxide (O 2 - ) production by thylakoids were investigated to determine if increased production of the radical was related to injury induced by chilling at a moderate photon flux density (PFD). Plants used were Spinacia oleracea L., Cucumis sativus L. and Nerium oleander L. grown at either 200° C or 45° C. Superoxide production was determined by electron-spin-resonance spectroscopy of the (O 2 - )-dependent rate of oxidation of 2-ethyl-1-hydroxy-2,5,5-trimethyl-3-oxazolidine (OXANOH) to the corresponding oxazolidinoxyl radical, OXANO ·. For all plants, the steady-state rate of O 2 - production by thylakoids, incubated at 25° C and 350 μmol photon · m−2 · s−1 (moderate PFD) with added ferredoxin and NADP, was between 7.5 and 12.5 μmol · (mg chlorophyll)−1 · h−1. Incubation at 5° C and a moderate PFD, decreased the rate of O 2 - production 40% and 15% by thylakoids from S. oleracea and 20° C-grown N. oleander, chillinginsensitive plants, but increased the rate by 56% and 5% by thylakoids from C. sativus and 45° C-grown N. oleander, chilling-sensitive plants. For all plants, the addition of either ferredoxin or methyl viologen increased the rate of O 2 - -production at 25° C by 75–100%. With these electron acceptors, lowering the temperature to 5° C caused only a slight decrease in O 2 - production. In the absence of added electron acceptors, thylakoids produced O 2 - at a rate which was about 45% greater than that when ferredoxin and NADP were present. The addition of 3-(3′,4′-dichlorophenyl)-1,1-dimethylurea reduced O 2 - production under all conditions tested. The results show that the rate of O 2 - production increases in thylakoids when the rate of electron transfer to NADP is reduced. This could explain differences in the susceptibility of thylakoids from chilling-sensitive and chilling-insensitive plants to chilling at a moderate PFD, and is consistent with the proposal that O 2 - production is involved in the injury leading to the inhibition of photosynthesis induced under these conditions.

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methyl viologen


Nerium oleander grown at 20° C


N. oleander grown at 45° C




photon flux density (photon fluence rate)


tetramethyl ethylenediamine


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We would like to thank R.T. Furbank, R.S.B.S., Australian National University, Canberra, A.C.T., and C.B. Osmond, now of Duke University, Durham, N.C., USA, for the gift of ferredoxin, R.A.J.H. was supported by a Commonwealth Postgraduate Research Award.

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Hodgson, R.A.J., Raison, J.K. Superoxide production by thylakoids during chilling and its implication in the susceptibility of plants to chilling-induced photoinhibition. Planta 183, 222–228 (1991). https://doi.org/10.1007/BF00197792

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Key words

  • Chilling
  • Cucumis
  • Nerium
  • Photoinhibition of photosynthesis
  • Photosynthesis (photoinhibition, active O2)
  • Spinacia
  • Superoxide