DETECTING STRESS-INDUCED REACTIVE OXYGEN SPECIES IN PLANTS UNDER UV STRESS - Environmental UV Radiation: Impact on Ecosystems and Human Health and Predictive Models - page 146

3.1 INDIRECT INDICATIONS
The occurrence of oxidative stress is often presumed from detecting damaged
products. In sugar beet leaves, UV-B irradiation was found to initiate lipid peroxidation
[15,16]. In isolated, photosynthetically active membrane preparations, UV-B results in
loss of electron transport activity and in fragmentation of reaction centre proteins D1
and D2 [17]. This is also observed in cyanobacteria, although a repair cycle based on de
novo protein synthesis may counterbalance damage in vivo (for review Ref. 18).
UV-B induced changes in the activity of antioxidant enzymes strongly suggests
the involvement of ROS. Although reports on the effect of UV-B irradiation on
antioxidants may vary with plant species, developmental stage and experimental
conditions, the general trend is activation of both enzymatic and non-enzymatic defence
(for review see Ref. 19). UV-B irradiation upregulated catalase and gluthatione
peroxidase in Nicotiana plumbaginifolia [20] induced most peroxidase-related enzymes
Arabidopsis [21], increased glutathione reductase transcript levels in Pisum sativum [22]
and Arabidopsis [23]. Reports on SOD are more controversial, ranging from little or no
[20,22] to marked activation [16,23]. UV-B was also found to induce pathogenesis-
related proteins in Arabidopsis [24]. The effect of UV-B is - in some aspects - similar
to other oxidative stresses. Plants respond to UV-B stress and pathogen infection
concerning both the free radical related ultraweak light emission [25] and the induction
of a stress response protein [26].
UV-B exposure increases the synthesis of UV absorbing pigments. [27]. Besides
UV screening, flavonoid compounds may also act as ROS scavenging antioxidants [28-
30]. Their central role is well illustrated by the increased susceptibility of barley [31]
and Arabidopsis [32,33] mutants with deficient flavonoid biosynthesis to UV-B
Increased levels of the free radical related ultraweak light emission upon UV-B
exposure of Hibiscus [34], sugar beet [15] and Brassica leaves [35] also indicated
oxidative stress.
3.2. DIRECT EVIDENCE
Although the paramagnetic nature of free radicals offers the possibility of
detection by electron paramagnetic resonance (EPR) spectroscopy, their high reactivity
and small concentration obstructs the application of this method. However, the use of
spin traps helps to overcome this difficulty making it possible to detect not only radical
natured ROS, but also singlet oxygen. As illustrated in Fig.4, spin traps are diamagnetic
(i.e. have no EPR signal), but form relatively stable, paramagnetic spin adducts when
reacting with ROS (for reviews see Refs. 36,37). Various spin traps are available
commercially.