The results obtained here support the applicability of the method to the fast and simultaneous determination of glutathione and glutathione disulfide in tissue of shoots and roots of plants grown under either normal or stressful conditions.
This study describes the fast and simultaneous determination of glutathione and glutathione disulfide by Capillary Zone Electrophoresis in plant extracts of shoot and root of tomato plants. Frequent use of acidic precipitation of protein generates an acidic matrix of strength and pH that may cause changes in the method sensitivity, comigration of species or changes in the equilibria that relate both species in cells or fluids. In this study, the resulting acidic matrix was previously treated with the same background electrolyte to prevent comigration and to improve signal resolution. Optimization of some parameters of the technique allowed the determination of both analytes in less than three minutes. The optimized method showed good reproducibility and linearity, with correlation coefficients above 0.999 and detection limits below 3 µM for both peptides. Analyte recovery in the process was in the 88-104% range. The concentration found in tomato plants hydroponically grown in the absence of stress factors was in the 51-100 nmol g -1 range, fresh weight for GSH and 5-32 nmol g -1 range, fresh weight for GSSG.
Phytotoxicity due to copper (Cu) is generally associated with visible symptoms such as a decrease in plant shoot and root growth and chlorosis. The application of low levels of Cu or plant exposure to the metal for short periods can affect several cell processes, involving changes in the levels of metabolites related to the plant antioxidative response. The purpose of this study was to compare the response of the reduced and oxidized glutathione system and Cu uptake in tomato plants exposed to phytotoxic levels of Cu in hydroponic culture. Attention was centered on establishing whether determination of the levels of both peptides in plants exposed for a short period of time (acute treatment) compared with exposure for an extended period (chronic treatment) could be used as an early indicator of Cu stress in tomato plants. For the acute treatment, the plants were exposed to 0.4, 9, and 36 μM Cu for 48 h, and for the chronic treatment to 0.4, 3, and 12 μM Cu for 28 d. Results indicate that plants subjected to the chronic treatment showed toxicity symptoms, among them chlorosis and a drastic decrease of the aerial part and root biomass, an effect that was not observed in the plants subjected to the acute treatment. Moreover, Cu applied to the plant, either in the acute or in the chronic treatment, modified the levels of reduced and oxidized glutathione in shoots and roots. The most noticeable effect was observed on the concentration of reduced glutathione in roots, where the concentration of this peptide decreased as the Cu concentration increased, and this effect was independent of the morphological changes undergone by the root and of the time of Cu application. The similarity of the responses to the acute and the chronic treatments indicates that assessment of the redox state of glutathione in the roots of plants exposed to copper, especially the change in levels of reduced glutathione, may represent a good indicator of the early plant response to stress due to excessive Cu supply.
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