Biochemical responses in leaves of two apple tree cultivars subjected to progressing drought.

The influence of water deprivation on potted apple trees (Malus domestica Borkh.) was investigated. Biochemical responses including ascorbic acid, glutathione (GSH), tocopherols, chlorophylls, carotenoids, free amino acids, and soluble carbohydrates were measured in leaves of diploid 'Elstar' and triploid 'Jonagold Wilmuta' subjected to progressive drought. Leaf water potential was chosen to be the primary indicator of water stress in tested plants. Time courses of measured biochemical parameters showed that mild drought did not significantly affect the chosen stress indicators. Moderate drought increased the concentrations of ascorbic acid, total GSH, beta-carotene, zeaxanthin and alpha-tocopherol, indicating the adaptation to oxidative stress in apple trees. Moderate drought also increased concentration of soluble carbohydrates, mostly due to increased sorbitol concentration. Severe drought negatively affected vitality of apple trees, and caused a decrease in sorbitol concentration. Severe drought also caused decreases in ascorbic acid, total GSH, beta-carotene, alpha-tocopherol and chlorophyll concentrations, which, together with the increase in oxidised GSH concentration, indicated severe damage due to oxidative stress. Severe drought increased free amino acid concentration, which was probably the result of increased proteolysis. Zeaxanthin concentration remained high even in leaves of apple trees subjected to severe drought stress. The results were similar for both tested apple cultivars.

Accumulating pollutants in conifer needles on an Atlantic island - a case study with Pinus canariensis on Tenerife, Canary Islands.

Concentrations of potential pollutant elements Na, Cl, and S were investigated in needles of Pinus canariensis grown at 55 field plots in Tenerife. Microelement concentrations (including heavy metals) were measured at a subset of 18 plots. Na and Cl concentrations were high at low elevations (up to 8 mg g(-1) Cl and 5.5 mg g(-1) Na). Na/Cl ratio close to standard seawater indicated sea spray influence up to 1200 m a.s.l. Only at few plots, sulphur concentrations indicated possible pollutant impact. Cluster and correlation analyses identified a related group of V, As, Cr, Fe, Mo, Ni, Cu, Pb, and Al, possibly related to traffic exhaust aggregated with soil particles. Mainly north-eastern, lower elevated plots were exposed to those immissions, but metal concentrations were generally low compared to data from other studies. In conclusion, seawater and soil particles explained most of the element distribution pattern in pine needles in Tenerife, but strong indications for some effect of local sources of air pollutants were detected.

Content of low-molecular-weight thiols during the imbibition of Pea seeds.

The metabolism of low-molecular-weight thiols was investigated in seeds of Pisum sativum L. cv. Kleine Rheinländerin during imbibition in water for 14 h. The amount of oxidized glutathione (GSSG) decreased from 319 nmol (g dry weight)-1 in dry seeds to 38 nmol (g dry weight)-1 within the first 14 h of imbibition. The decrease may have been due to the reduction of GSSG to reduced glutathione (GSH), catalyzed by the enzyme glutathione reductase (GR; EC 1.6.4.2). The enzyme activity was high in dry seeds [25 nkat (g dry weight)-1 ] and decreased to 20 nkat (g dry weight)-1 within 14 h of imbibition. The activity of glucose-6-phosphate dehydrogenase (EC 1.1.1.49) decreased from 100 nkat (g dry weight)-1 in dry seeds to 67 nkat (g dry weight)-1 after 14 h of imbibition. Within 14 h the amount of γ-glutamyl-cysteine (γ-GC) decreased from 135 to 38 nmol (g dry weight)-1 , whereas the cysteine content rose from 81 nmol (g dry weight)-1 in dry seeds to a maximum of 170 nmol (g dry weight)-1 after 12 h of imbibition, which may be due to the degradation of γ-GC into cysteine.

Root uptake, transport, and metabolism of externally applied glutathione in Phaseolus vulgaris seedlings.

The most abundant thiol in beans (Phaseolus vulgaris L. cv. Saxa) is the tripeptide homoglutathione (hGSH) rather than glutathione (GSH). At the whole-plant level the GSH content is less than 0.5% of the hGSH content. In the present study GSH was supplied to the roots of bean seedlings to test whether GSH can be taken up by roots and transported to the shoot. Therefore, 12-day-old plants were exposed to 1 mmol/L GSH for 4, 8 and 24 h prior to harvest. In response to this GSH exposure, elevated GSH contents were found in all tissues. After 4 h the GSH content increased in the roots from 1 +/- 1 to 22 +/- 2 nmol GSH g(-1) fresh weight (FW), in the leaves from 2 +/- 1 to 9 +/- 4 nmol GSH g(-1) FW, and in the apex from 30 +/- 5 to 75 +/- 4 nmol GSH g(-1) FW. These data indicate that GSH is taken up by bean roots and is transported to above above-ground parts of the plants. Roots exposed to GSH for 24 h contained 2-fold higher cysteine (Cys) and hGSH contents than the controls. Apparently, GSH taken up by the roots is not only loaded into the xylem but also partially degraded and used for hGSH synthesis.

Multivariate patterns of antioxidative and photoprotective defence compounds in spruce needles at two central European forest sites of different elevation.

In this work we measured a set of antioxidative and photoprotective compounds (chlorophylls, carotenoids, tocopherol, ascorbate and glutathione), which were suggested previously as stress markers in conifer needles, at two spruce forest sites at different elevation in Saxony, Germany. Most variables differed significantly between current and 1-year-old needles, but only the content of the xanthophyll cycle per mg total chlorophyll and the oxidation state of glutathione were significantly different between the sites. We applied principal component analysis (PCA) to address the question if underlying accumulated variables are similar to the ones found in spruce needles across Alpine elevation profiles and/or for pines in Mediterranean ecosystems. Four principal components (accumulated variables, PC) representing 68% of the total variance of the dataset were extracted. PC 1 encompassed total chlorophyll, lutein, and beta-carotene contents, PC 2 combined the epoxidation state of xanthophylls, ascorbate content and redox state, and glutathione content, PC 3 represented the content of xanthophylls and the redox state of glutathione, and PC 4 encompassed the content of alpha-carotene and the epoxidation state of xanthophylls. Only PC 3 was significantly different between sites. The PCA structure shows many similarities to corresponding findings in studies on spruce in mountain forests in the Alps and pines in Mediterranean systems. This corroborates the interpretation of PCs as indicative for underlying physiological processes. However, separation of the two investigated sites by PCs was in the present case study not superior to the separation by single input variables.

The glutathione system as a stress marker in plant ecophysiology: is a stress-response concept valid?

Environmental stress impacts cause an increased formation of reactive oxygen species (ROS) in the chloroplasts (photo-oxidative stress). The role of glutathione in the antioxidative defence system provides a rationale for its use as a stress marker. However, responses of glutathione concentrations and redox states are not consistent among the large number of available publications. In the present review the hypothesis that stress responses of the glutathione system follow a general ecophysiological stress-response concept is investigated. In this view, an initial response phase would be followed by an acclimation phase where a new steady-state is established. Alternatively, if successful acclimation is not achieved, degradation of the system will follow. Recent publications dealing with responses to photochilling, salinity, and drought are analysed as to whether the results fit the concept. In general, an initial stress response was related to changes in the glutathione redox state, whereas acclimation was marked by increased glutathione concentrations, increased related enzyme activities, and/or a more reduced redox state of glutathione. The latter was interpreted as overcompensation leading to enhanced regeneration of glutathione. Deterioration effects upon strong stress impacts were related to progressive degradation and oxidation of the glutathione pool. A time-course analysis, which has rarely been done in the published literature, showed this sequence of events. When apple trees were subjected to progressing drought, the initial response was a slight oxidation of the glutathione pool, followed by increased glutathione concentrations. When the stress increased, glutathione concentrations dropped and redox state became more oxidized, which marked the degradation of the system. In spite of the general congruency of these results with the suggested stress-response concept, several limitations have to be highlighted: The importance of the glutathione system relative to other components of the photoprotective and antioxidative defence system, as well as relative to stress avoidance strategies, has to be established. It is suggested that a variety of parameters taking into account alternative protection pathways (e.g. photorespiration, light dissipation) and other components of the antioxidative systems should be measured. Within such response patterns the glutathione system is a valuable stress marker in ecophysiological studies.

Effects of weak 16 3/2 Hz magnetic fields on growth parameters of young sunflower and wheat seedlings.

Previous studies on effects of magnetic fields on plants focussed on the power frequencies (50-60 Hz), but in Austria and Germany electric railways are powered by a frequency of 16 2/3) Hz. In the present study, sunflower and wheat seedlings were exposed to 16 2/3 Hz sinusoidal 20 muT (rms) vertical magnetic fields. Seeds were germinated in a germination roll and grown for 12 days under continued exposure. Seven series with sunflower and six series with wheat were done over 2 years. Sunflower seedlings exposed to experimental magnetic field showed small, but significant increases in total fresh weights, shoot fresh weights, and root fresh weights, whereas dry weights and germination rates remained unaffected. Experimentally treated wheat exhibited marginally (but significantly) higher root fresh and dry weights, total fresh weights, and higher germination rates.

Measuring antioxidants in tree species in the natural environment: from sampling to data evaluation.

Biochemical measurements of antioxidants and protective pigments have been successfully introduced as markers of environmental stress in field studies (mainly forest studies). A guideline for field sampling and analysis methods is required to allow better comparison of data from different studies. The present review paper recommends HPLC methods for the analysis of ascorbate and glutathione (in oxidized and reduced form), tocopherols, and chloroplast pigments. Methodological variations are substantially lower (coefficients of variance of repeated extractions typically 4-9%) than biological variations of field samples (typical variation coefficients 8-36%), hence special emphasis is put on considerations of sampling standardization in the field with respect to sample time (seasonal and diurnal) and representative sampling of individuals and tissues. Following the suggestions in this paper would enable researchers to produce results that could be compared with those of several forest studies on conifers published in recent years. A larger data-set available for multivariate statistical evaluations (e.g. principal component analysis and cluster analysis) will enhance the diagnostic value of such investigations.