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Widespread use of silver nanoparticles (Ag NPs) as pesticides and fungicides in agriculture is a major environmental concern. In the present study, soybean (Glycine maxL.) was grown in Ag NP (0–62.5 mg kg−1)-amended soil with or without the addition of glutathione (GSH). Ag NPs exerted severe phytotoxicity and caused the reduction of shoot and root biomass and total number of nodules or completely inhibited nodule formation at doses above 31.25 mg kg−1. Synchrotron-based techniques were applied to analyze Ag speciation in both the soil and the soybean root tissues at harvest. The results indicate that the majority of Ag remained in the form of Ag NPs and that 23% was present as Ag2S in soil; in both root and nodule tissues, Ag–GSH was the main component (40.6–88%) other than Ag NPs (12–59.4%), highlighting the important role of GSH in alleviating the Ag NP-induced toxicity. The addition of 0.8 mM GSH not only significantly increased fresh biomass by 85% in the 62.5 mg kg−1Ag NP treatment but also decreased Ag accumulation by 24.8–27% in soybean tissues. Although the addition of 0.8 mM GSH reduced the nodule number and weight as compared to the control, the total nitrogen content in soybean co-treated with Ag NP and GSH was more than 5-fold higher than that in the Ag NP alone treatments, suggesting that GSH may be utilized as a nitrogen source while simultaneously alleviating Ag NP toxicity. The shoot and root contents of thiol compounds (cysteine and gamma-glutamylcysteine) in the GSH treatments were several folds higher than that in the control and Ag NP alone treatments. Further, higher levels of essential amino acids, particularly alanine, glutamate, and glutamine which play important role in N assimilation in plants, in soybean across all the treatments further confirmed that GSH was utilized as a nitrogen source, resulting in enhanced soybean growth. Taken together, this study clearly demonstrated the negative impact of Ag NPs on soybean productivity and N fixation and highlights the protective role of GSH against Ag NP-induced toxicity. These findings have significant relevance for developing future strategies to minimize crop loss in marginal or contaminated soils, subsequently enhancing global food security.

Abstract Tree care, especially pruning, is still mostly done in the dormant time. Such treatments expose live inner bark, the vascular cambium, and functioning outer sapwood to harsh external influences followed often by infection of pathogens. Investigations about response reactions of beech and oak to wounding made in different times of the year showed that wound closure was significantly slower for December and February wounds than in April and October. The length of discoloured sapwood was in October and December wounds greater than in February and April. Sapwood reaction expressed by the soluble phenol concentration index indicated however no significant differences among wounding dates. Overall, woundings in the vegetation period will be more effectively compartmentalized than in the dormant season. Consequently, tree care like pruning with many woundings should not be done in the winter period.

Summary??The major objective of this study was to determine if the observed changes in polyamines and their biosynthetic enzymes during somatic embryo development were specifically related to either the stage of the embryo development or to the duration of time spent on the maturation medium. Somatic embryos of red spruce (Picea rubens) at different developmental stages, grown in the embryo development and maturation media for various lengths of time, were separated from the associated subtending tissue (embryogenic and the suspensor cell masses) and analyzed for their polyamine content as well as for polyamine biosynthetic enzyme activities. Polyamine content was also analyzed in embryos representing different stages of developmentthat were collected from the sam culture plate at the same time and the subtending tissue surrouding them. Putrescine was the predominant polyamine in the pro-embryogenic tissue, while spermidine was predominant during embryo development. Significant changes in spermidine/putrescine and spermine/putrescine ratios were observed at all stages of embryo development as compared to the pro-embryogenic cell mass. Changes in the ratios of various polyamines were clearly correlated with the developmental stage of the embryo rather than the period of growth in the maturation medium. Whereas the activities of both ornithine decarboxylase and arginine decarboxylase increased by week 3 or 4 and stayed high during the subsequent 6 wk of growth, the activity ofS-adenosylmethionine decarboxylase steadily declined during embryo development.

We investigated the catabolism of putrescine (Put) in a non-transgenic (NT) and a transgenic cell line of poplar (Populus nigra x maximowiczii) expressing a mouse (Mus musculus) ornithine (Orn) decarboxylase (odc) cDNA. The transgenic cells produce 3- to 4-fold higher amounts of Put than the NT cells. The rate of loss of Put from the cells and the initial half-life of cellular Put were determined by feeding the cells with [U-(14)C]Orn and [1,4-(14)C]Put as precursors and following the loss of [(14)C]Put in the cells at various times after transfer to label-free medium. The amount of Put converted into spermidine as well as the loss of Put per gram fresh weight were significantly higher in the transgenic cells than the NT cells. The initial half-life of exogenously supplied [(14)C]Put was not significantly different in the two cell lines. The activity of diamine oxidase, the major enzyme involved in Put catabolism, was comparable in the two cell lines even though the Put content of the transgenic cells was severalfold higher than the NT cells. It is concluded that in poplar cells: (a) exogenously supplied Orn enters the cells and is rapidly converted into Put, (b) the rate of Put catabolism is proportional to the rate of its biosynthesis, and (c) the increased Put degradation occurs without significant changes in the activity of diamine oxidase.

Traditionally, most enzyme assays utilize homogenized cell extracts with or without dialysis. Homogenization and centrifugation of large numbers of samples for screening of mutants and transgenic cell lines is quite cumbersome and generally requires sufficiently large amounts (hundreds of milligrams) of tissue. However, in situations where the tissue is available in small quantities, or one needs to study changes in enzyme activities during development (e.g. somatic embryogenesis), it is desirable to have rapid and reproducible assay methods that utilize only a few milligrams of tissue and can be conducted without homogenization. Here, we report a procedure for the measurement of enzyme activities of the three key decarboxylases involved in polyamine biosynthesis utilizing small quantities of plant tissue without the homogenization and centrifugation steps. Suspension cultures of red spruce (Picea rubens(Sarg.)), hybrid poplar (Populus nigra× maximowiczii), and wild carrot (Daucus carota) were used directly to measure decarboxylation of ornithine, arginine and S-adenosylmethionine. Our results demonstrate that this procedure can be used to quantify the activities of arginine decarboxylase (EC 4.1.1.19), ornithine decarboxylase (EC 4.1.1.17) and S-adenosylmethionine decarboxylase (EC 4.1.1.50) in a manner quite comparable to the traditional assays for these enzymes that involve laborious steps of homogenization and centrifugation.

The effects of aluminum (Al) treatment on polyamines were studied using suspension cultures of Madagascar periwinkle [Catharanthus roseus(L.) G. Don]. The addition of Al (0.2, 0.5, 1.0 mM) to the suspension cultures caused a significant increase in putrescine within 24 h only in freshly transferred cells. By contrast, Al treatment reduced putrescine content at all other times (days 2–7). Further analysis showed that the largest increase in putrescine occurred at 6 h when Al was added at the time of transfer of cells to fresh medium. If Al treatment was delayed to as little as 3 h after transfer to fresh medium, the cellular level of putrescine decreased. While spermidine remained unchanged, spermine was significantly higher in the presence of Al in all cases. The inhibitory effect of Al on DNA synthesis also depended upon the time of its addition. Cellular Ca, Mg, and Mn decreased in response to Al given at the time of transfer to fresh media while Fe increased, and Na, K and P remained unchanged. The results indicate a critical physiological change in the response of cells to Al within a short time (about 3 h) after transfer to fresh medium.

Dendrochemical and biochemical markers link stress in apparently healthy red spruce trees (Picea rubens) to acidic deposition. Acidic deposition to spruce forests of the northeastern USA increased sharply during the 1960s. Previous reports related visible damage of trees at high elevations to root and soil processes. In this report, dendrochemical and foliar biochemical markers indicate perturbations in biological processes in healthy red spruce trees across the northeastern USA. Previous research on the dendrochemistry of red spruce stemwood indicated that under uniform environmental conditions, stemwood concentrations of Ca and Mg decreased with increasing radial distance from the pith. For nine forest locations, frequency analysis shows that 28 and 52% of samples of red spruce stemwood formed in the 1960s are enriched in Ca and Mg, respectively, relative to wood formed prior to and after the 1960s. This enrichment in trees throughout the northeastern USA may be interpretable as a signal of increased availability of essential cations in forest soils. Such a temporary increase in the availability of Ca and Mg could be caused by cation mobilization, a consequence of increased acidic deposition. During cation mobilization, essential Ca and Mg as well as potentially harmful Al become more available for interaction with binding sites in the soil and absorbing roots. As conditions which favor cation mobilization continue, Ca and Mg can be leached or displaced from the soil. A measure of the interaction between Ca and Al is the Al/Ca binding ratio (molar charge ratio of exchangeable Al to exchangeable Ca). As the Al/Ca binding ratio in the root zone increased from 0.3 to 1.9, the foliar concentration of the biochemical stress marker putrescine also increased from 45 to 145 nm g−1. The correlation of the putrescine concentration to the Al/Ca binding ratio (adj. r2= 0.68, P< 0.027) suggests that foliar stress may be linked to soil chemistry.

A fast and reliable method for the extraction of cellular polyamines and major inorganic ions (Ca, Mg, Mn, K, and P) from several plant tissues is described. The method involves repeated freezing and thawing of samples instead of homogenization. The efficiency of extraction of both the polyamines and inorganic ions by these two methods was compared for 10 different tissues. In each case, the freeze-thaw procedure resulted in a precise and quantitatively equal, or greater, yield than homogenization. Freeze-thawing not only eliminates the need for various tissue homogenizers (such as polytrons, tissumizers, and mortars and pestles), but it is so simple that a large number of samples can be processed simultaneously. We routinely processed 50–80 samples for quantitation of polyamines and inorganic ions. Freeze-thawing was equally useful for the extraction of polyamines from liver, spleen, and kidney tissues of mice.

This paper describes a fast reliable, and a sensitive technique for the separation and quantification of dansylated polyamines by high-performance liquid chromatography. Using a small 33 × 4.6 mm I.D., 3 μm particle size, C18reversed-phase cartridge column and a linear gradient of acetonitrile—heptanesulfonate (10 mM, Ph 3.4), at a flow-rate of 2.5 ml/min, the retention time for different polyamines was: N8-acetyl-spermidine, 1.79 min; N1-acetylspermidine, 1.82 min;putrescine, 2.26 min; cadaverine, 2.43 min; heptanediamine, 2.83 min; spermidine, 3.42 min; and spermine, 4.41 min. With an additional column regeneration time of 3—4 min, the complete cycle per sample took less than 8 min at room temperture. Using a fluorescence detector, the lower limit of detection was less than 1 pmol per 6 μl injection volume. The fluorescence response was linear up to 200 pmol per 6 μl for each polyamine. The method is suitable for separation of polyamines from animal, plant and fungal sources.