O in bread wheat Triticum

Ascorbate peroxidase (APX, EC 1.11.1.11) 
activity assay. The activity of the enzyme was 
determined spectrophotometrically based on the 
decomposition of H
2
O
2 
by the ascorbate peroxidase 
enzyme for 1 min, at 290 nm (Nakano and Asada, 
1981). The reaction medium consisted of 0.1 mM 
EDTA (pH 8.0), 0.05 mM ascorbic acid, 0.1mM 
H
2
O
2
, 50 mM Na-Phosphate (pH 7.6) buffer and 
100 µl of the enzymatic extract. The APX activity 
was estimated based on the decline in the optic 
density during the first 30 sec of the reaction and 
was expressed in mmol ascorbate/(mg protein min) 
at the coefficient of extinction (ɛ) of 2.8 mM
-1
cm
-1
. 
Glutathione reductase (GR, EC 1.6.4.2) 
activity assay. The enzyme activity was determined 
spectrophotometrically at 340 nm in the presence of 
oxidized glutathione (GSSH), based on the 
oxidation of NADPH for 3 min. Glutathione 
Reductase Assay Kit (Sigma-Aldrich) was used for 
this purpose. The enzyme activity was expressed in 
µmol/(mg protein min) and the molar extinction 
coefficient was assumed to be ε=6.2 mM
-1
cm
-1
. 
Total protein content determination was 
based on the method of Bradford (1976). Bovine 
serum albumin was used to construct the calibration 
curve. 
Statistical analysis: In all figures, values are 
shown as error bars representing standard errors of 
the means. The significance of differences between 
mean values was compared by Student’s T-test.
 
 
RESULTS AND DISCUSSION 
 
As under field conditions, plants are exposed 
to various stresses at the same time, the AsA-GSH 
cycle plays an important role in the regulation of 
defense responses to the destructive oxidizing 
effects of both pathogens and abiotic stresses 
(Elzbieta et al., 2017). Although numerous reports 
are confirming an important role of ascorbate and 
glutathione in the prevention of oxidative stress, 
there is still little information on this area 
(Hasanuzzaman et al., 2019). The presence of the 

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L.M. Aydinli 
64 
AsA-GSH components and the detected activities 
of catalase and superoxide dismutase in the tolerant 
Amaranthus tricolor variety show that these 
substances play an important role in scavenging 
ROS from plants (Sarker and Oba, 2018). 
Leaf samples were taken in the last decade of 
May in 2017, 2018, 2019 from 3 variants (control, 
drought-exposed, rehydrated) of the plants grown 
under field conditions, and the following results 
were obtained: The amount of AsA increased in the 
Gobustan variety by 91% under drought and 
recovered by 70% after rehydration. In the Tale 38 
variety, the AsA amount increased by 79% in the 
drought-exposed variants and recovered only by 
15% after rehydration (Fig. 1A). The ascorbate 
peroxidase activity increased by 13% in the 
drought-exposed Gobustan variety and recovered 
after rehydration only by 1.5%. According to the 
results of the statistical analysis, there is no 
significant difference between the control and 
rehydrated variants. This indicates the high level of 
the recovery processes in the Gobustan variety. In 
the Tale 38 variety, the APX activity decreased by 
42% in the drought-exposed plants compared with 
the control variant and a 22% decrease compared 
with the control variant was observed after 
rehydration (Fig. 1B). 
Endogenous 
accumulation 
of 
reduced 
glutathione in plants was found to increase plant 
tolerance to drought and decrease the harmful 
effects of ROS (Hasanuzzaman et al., 2017). The 
changes in the glutathione amount accumulated in 
leaves of the plants at the wax ripening stage of 
ontogenesis are presented in Figure 2 (A, B). In the 
drought-exposed plants of the Gobustan variety 
GSH amounts on a fresh weight basis increased by 
33%, and recovered by 16% after rehydration. In 
leaves of the drought-exposed variants of the Tale 
38 variety, accumulation of reduced glutathione 
increased by 17% and recovery in the rehydration 
variants was 5% (Fig. 2B).