Membrane Lipid Peroxidation and Plasma Membrane Permeability
To explore the relationship among the leaf aging, lipid peroxidation, cell signaling, and different light environments, reactive oxygen species (Superoxide-O2.– and Peroxide-H2O2 radicals) and malondialdehyde (MDA) was quantified under given conditions at 1, 10, 20, 30, and 40 days after flowering initiation. Quantitative analysis of reactive oxygen species revealed that with increasing leaf age, H2O2 and O2.– contents increased under all light treatment conditions. Initially, PL significantly induced ROS production during the first ten days, particularly in leaves from the shaded side (PS) compared to PF, NF, and NS (Fig. 2B-C). However, during subsequent days (20-40), the situation reversed, and the maximum production of H2O2 and O2 was noticed under NL as compared to the minimum production of ROS under PS conditions. Notably, there was no significant difference observed between NF and NS regarding ROS production. Within PL, the PF condition produced significantly more ROS (14.2% H2O2, 22.3% O2.–, 6.0% MDA) than PS, except for the initial ten days of treatment when both treatments behaved the same (Fig. 2B-C). A similar trend was visualized by histochemical staining (Fig. 2A-C). Furthermore, the increased amount of MDA showed that lipid peroxidation followed almost the same trend as ROS production under NF, NS, and PF. Contrarily, the MDA production under PS conditions increased to the maximum 37.6 μmol g-1 for the first 20 days and then declined significantly to the minimum 30.8 μmol g-1 compared to NF, NS, and PF (Fig. 2D).
To investigate the difference in ROS detoxification, we analyzed the activity of the three most critical antioxidant enzymes (SOD, POD, CAT) that scavenge excessive ROS. The results showed a highly significant increase in the activity of SOD for the first 20 days and POD and CAT for the first ten days under PS after that it started to decline (Fig. 2E-G). On the other hand, there were no significant differences observed between NF and NS despite the increase in activity (POD, CAT up to 30 days; SOD up to 20 days).