Effects of β - Carotene & Gallic Acid Pre-Treatment on Seedlings Total Fresh Weight Under Salt Stress
According to the findings of preliminary experiments to determine the concentration of sodium chloride, β-carotene, and gallic acid: 1- for salinity treatment, 25 mM sodium chloride caused a minimal reduction in seedling growth indices of L. sativum seedlings (Data not shown, Figure Suppl number 1-5). 2- Concentration of 0.5 mM β-carotene caused a maximum significant increase in growth indices of L. sativum seedlings (Data not shown, Figure Suppl number 6-9). 3- 5 mM gallic acid concentration caused the maximum significant increase in seedling growth indices of L. sativum (Data not shown, Figure Suppl number 10-13). Therefore, 0.5 mM β-carotene and 5 mM gallic acid were used to investigate the reduction of the effects of 25 mM sodium chloride in L. sativum seedlings.
Compared to sodium chloride treatment, according to Figure 2A, β-carotene and gallic acid pretreatment increased the total fresh weight by 44% and 40%, respectively. Salinity stress resulted in a 28% reduction in total fresh weight compared to control. The total fresh weight of seedlings treated with β-carotene + sodium chloride increased by 37% compared to seedlings treated with sodium chloride.
Figure 1B showed that β-carotene alone did not significantly change the total dry weight compared to gallic acid alone, but both led to an increase in total dry weight compared to control. Sodium chloride treatment resulted in a 22% reduction in total dry weight compared to the control. β-carotene + sodium chloride treatment resulted in a 48% increase in total dry weight compared to sodium chloride. Total dry weight under gallic acid + sodium chloride treatment increased by 27% compared to sodium chloride. Comparison of mean data showed that the highest total dry weight was observed in seedlings treated with β-carotene + sodium chloride and β-carotene alone, and the lowest root fresh weight was observed in seedlings treated with sodium chloride.
Effects of β - Carotene & Gallic Acid Pre-Treatment on Total Chlorophyll Content Under Salt Stress
Salinity treatment had a negative effect on total chlorophyll content, while pretreatment of β-carotene and gallic acid increased the content of total chlorophyll compared to the control. An increase in the total chlorophyll content of 35% and 44% was observed in β-carotene + NaCl and gallic acid + NaCl treatments in comparison to the NaCl treatment, respectively (Figures 3).
Effects of β - Carotene & Gallic Acid Pre-Treatment on H 2 O 2 & Malondialdehyde (MDA) Content Under Salt Stress
The H2O2 content reported in figure 4A, the results showed that the H2O2 content under β-carotene and gallic acid pre-treatment decreased by 13% and 17%, respectively, compared to the control. Sodium chloride treatment increased H2O2 by 63% compared to the control. β-carotene + sodium chloride and gallic acid + sodium chloride treatments reduced H2O2 by 33% and 37% compared to sodium chloride, respectively. As shown in figure 3B, the amount of MDA in treatment with β-carotene, gallic acid decreased by 6% and 5% compared to the control. Sodium chloride treatment showed a 99% increase in MDA compared to the control. β-carotene + sodium chloride and gallic acid + sodium chloride treatments reduced MDA content by 19% and 22%, respectively, compared to sodium chloride.
Effects of β - Carotene & Gallic Acid Pre-Treatment on Phenolic, Glutathione Content & DPPH Radical Scavenging Activity Under Salt Stress
The findings of figure 5A revealed that the content of phenolic compounds in β-carotene, gallic acid, and sodium chloride treatments increased by 15%, 19%, and 11%, respectively, compared to the control. β-carotene + sodium chloride and gallic acid + sodium chloride treatments increased phenolic compounds by 28% and 15%, respectively, compared to sodium chloride.
The results of figure 5B showed that the antioxidant activity of the plant under the treatments of β-carotene, gallic acid, and sodium chloride increased by 44%, 46%, and 38%, respectively, compared to the control. β-carotene + sodium chloride and gallic acid+sodium chloride treatments increased the plant's antioxidant activity by 4% and 6%, respectively, compared to sodium chloride.
The findings of figure 5C demonstrated that the content of glutathione in β-carotene and gallic acid pre-treatment increased by 56% and 37%, respectively, compared to the control. In sodium chloride treatment, a 3.84-fold increase in the amount of glutathione was observed compared to the control. The amount of glutathione under β-carotene + sodium chloride and gallic acid + sodium chloride treatments decreased by 38% and 52%, respectively, compared to sodium chloride.
Effects of β - Carotene & Gallic Acid Pre-Treatment on Antioxidant Enzymes Under Salt Stress
Consistent with the findings, CAT, GPX, APX, SOD, and GR enzymes activities increased in the salinity-treated plants compared with the control. β-carotene and gallic acid pretreatments alone significantly increased the activities of CAT, GPX, APX, SOD, and GR enzymes compared to the control. A relatively larger increase in the activities of CAT, SOD, GR was observed in gallic acid treatment as compared to β-carotene. β-carotene + sodium chloride and gallic acid + sodium chloride treatments significantly increased the activities of CAT, GPX, APX, SOD, and GR enzymes compared to sodium chloride; however, the increase in the activities of SOD, GPX was more in gallic acid treatment than the β-carotene treatment under salinity stress. While the use of gallic acid and β-carotene in the activities of GPX and CAT was not significantly different under salinity conditions compared to each other (Figure 6A, B, C, D).
Effects of β - Carotene & Gallic Acid Pre-Treatment on Ions Analysis Under Salt Stress
Salinity and β-carotene, gallic acid treatments showed a significant interaction on Na+, K+ contents, K+/Na+ ratio of shoot and root. Salinity stress reduced the K + content, K+/Na+ ratio but increased Na+ accumulation of roots and shoots compared to the control. Application of β-carotene alone and gallic acid alone significantly increased K+ contents, K+/Na+ ratio of shoot and root but decreased Na+ content. In shoot and root, β-carotene showed more effect on Na+ reduction and K+ content, K+/Na+ ratio improvement compared with gallic acid. Treatment of β -carotene, gallic acid under salinity stress improved K+ content and K+/Na+ ratio of roots and shoots to stress conditions and reduced Na+ uptake, indicating the positive effect of this pretreatment on increasing K+ ions and decreasing Na+ (Figure 7A, B, C, D, E, F).