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Well-established label in biomolecules analysis, fluorescein isothiocyanate is commercially available in its two different isomers with equal fluorescence sensitivity, differing in meta- vs. para-isothiocyanate position. It was found in this study that the stability of the isomers depends on the polarity of different solvents and the pyridine (stabiliser) presence. The lowest stability has fluorescein isothiocyanate in water solvent. In acidic solution, their quinoid (acid) form switches to the lactone form. The p-quinoid form has higher tendency to create dimers and trimers and is less soluble in acetone than the lactone form. [GRAPHICS] .

Triazoles are used as antifungal agents, they mostly inhibit two enzymes: 14α-demethylase and aromatase. These enzymes are utilised also in other species and therefore the affection in non-target species in the environment is expected as well. Besides, triazoles are often being applied in a mixture and they can also interact with other substances present. This study clarifies how three selected representative triazoles (tebuconazole, penconazole and cyproconazole) interact with each other (group effect) and in mixtures (cocktail effect) with copper, essential/toxic for all organisms. Within the experiments on electrospray and collision-induced dissociations (both ESI-MS), it has been found that the fragments correspond to typical triazole metabolites. For their formation, the presence of copper ions is crucial. The inhibitory effect of Cu cocktails on aromatase enzymatic activity has been studied. The presence of Cu ions together with triazole(s) significantly increases the inhibitory effect on aromatase activity. The highest inhibitory effect (more than 60%) on aromatase activity is produced by cocktails containing penconazole and Cu ions, namely by penconazole/Cu and penconazole/tebuconazole/Cu. The reactivity of triazoles in groups is not significantly affected by the interactions among them. Additionally, the role of triazoles in copper Fenton reaction regulation has been observed and described. These changes may be attributed to the formation and stabilization of the complexes with the central Cu ion, with usually one, two or three triazolic ligands, depending on the mixture. The study demonstrates that the interaction of triazoles and Cu ions is a complex process; their impact on metabolism seems to be rather extensive and must be evaluated in the context of biochemical reactions.

Triazolic fungicides are widely applied in crop production to protect plants against fungal pathogens. However, they may influence the biochemical processes in plants and other non-target species. This paper is aimed at the effect of triazoles (namely tebuconazole, cyproconazole, and penconazole) single/mixed applications on the phenolics production in tomato (Solanum lycopersicum L.) fruit peel, amount of chlorophyll a and b in tomato leaves as well as on basic plant growth parameters. For this purpose, cherry tomatoes were planted in the pot experiment and foliarly-treated weekly, with the same total triazoles dose of 3.52 μmol per plant (in mixtures of 1.71 or 1.17 μmol of each in two or three components, respectively). The treatments increased the weight of fruits in the 1 harvest about 43%, however, this effect was not observed in the next harvest. Increased oxidative stress in the triazoles presence was observed, based on the elevated production of antioxidant phenolics in the 1 harvest. Most alarming is the decrease of the weight of thin stems and foliage and the concentration of chlorophyll a (b) in leaves in all triazoles-treated variants. The non-target impacts on plant biochemical processes (related to the phenolics or chlorophylls production and functionality) were confirmed.

The azoles (represented by penconazole, cyproconazole, and tebuconazole in this study) are frequently used agrochemicals to protect various crops against mildew and fungi. They are considered as endocrine disruptors, because they block the biosynthesis (on the level of enzymes inhibition) of biochemicals with steroid structure. Besides targeted impacts, they can partly get into the soil with the rainfall or litter fall and influence/block the biosynthesis of sterols of non-target organisms. In this sense, the risk of disruption of rhizosphere plant-microbial symbiosis and dynamic processes in the soil solution by azoles is of high importance to be evaluated. We have developed an analytical methodology for determination of penconazole, cyproconazole, and tebuconazole in soil solution using capillary electrophoresis with a photodiode array detector at UV-214 nm and acidic electrolyte solution (pH 1.48). The results were also compared with mass spectrometric measurements using mu-TOF mass spectrometry. There approx. 90% of present azoles were bound in the soil solution matrix. The detection limit for these azoles is about 10(-7) mol dm(-3). Because of very low pK(a) of azoles, we have to consider deprotonation of azoles and consequently the high affinity to create complexes with cations. The majority of present azoles in soil solution might form neutral adducts with mono-cations, making them invisible in electrospray mass spectra. [GRAPHICS] .

Dimethyl sulfoxide (DMSO) is widely utilized compound in biomedical and technological applications. Predominantly, it is used for biomembranes opening during a drug delivery procedure. The mechanism of DMSO membrane treatment is influenced by DMSO interactions with Ca 2+ as well as with water presence. In this study we investigate Ca 2+ interactions with DMSO in an aqueous environment using electrospray ionization mass spectrometry supported by density functional calculations. The number of DMSO ligands in [Ca(DMSO) x ] 2+ depends on the conditions during the electrospray and ion-transfer process. At capillary temperatures lower than 120 °C the complexes with a high number of DMSO are preferentially transferred to the gas phase (e.g. [Ca(DMSO) 6 ] 2+ ). With increasing temperature the number of DMSO ligands decreases while they are replaced with water molecules. Both the experiment and the calculations show that the bonding energy of DMSO to Ca 2+ is strongly influenced by the presence of water. The calculations also demonstrate that the bonding of DMSO to Ca 2+ is rather strong; the Ca 2+ -DMSO bonding energy was calculated to be two times higher than the Ca-H 2 O bonding energy.

The frequent use of pesticides in crop protection leads to their accumulation in soil as well as to the formation of their complexes with ions present in soil. The formation of such complexes can change the mobility of both the pesticides and the ions as well as their ability to penetrate biological membranes and enter biota. The problem of triazole-fungicide complexation with dications is demonstrated on the case of zinc and cadmium ions (M). Complexation with tebuconazole ( 1 ) has been studied using electrospray ionization mass spectrometry and quantum mechanical DFT-based computations. These experiments have revealed that the deprotonation of 1 leads to the stabilization of the complexes formed, which has been confirmed by our computations. In terms of stability, two types of complexes have been described: (i) a less stable complex (approx. 125 kJ mol −1 ) with neutral 1 [M( 1 ) 2,3,4 ] 2+ and [MCl( 1 ) 1,2 ] + , and (ii) a more stable complex (approx. 220 kJ mol −1 ) with deprotonated 1 [M( 1 ) n ( 1 -H)] + ( n = 0,1). The high stability of the complexes with deprotonated 1 is given by their stabilization via a metal interaction with the oxygen atom or phenyl ring. The structure of [M( 1 -H)] + is very compact when compared to the structure of parent 1 .

Tebuconazole ( 1 ) belongs to a class of widely utilized plant fungicides, mostly in vineyards. Consequently, traces of this compound are present in plant tissues, wine as well as in soil. The aim of this study was in situ generation of Cu cations and creation of very stable Cu complexes with 1 . The proposed method involves a hyphenation of electrochemistry with electrospray ionization mass spectrometry (ESI-MS), based on the high affinity of 1 for dissolved copper cations. We demonstrate that at appropriate potentials (above 500 mV vs. silver reference electrode) Cu cations can be generated in an electrochemical cell, which subsequently form complexes with 1 present in the sample. The resulting complex [Cu( 1 )] + as well as its fragment [Cu( 1 -C 4 H 10 )] + are then detected with online-coupled ESI-MS. This finding is useful for a creation of an analytical method based on the detection of trace amounts of 1 in complex matrices (e.g., soil solution), in which the analyte may be spread among various species.

A young speckled alder ( Alnus incana (L.) Moench) stand was planted on a tract clear-felled due to air pollution and located on a summit plateau of the Jizerske hory Mts. (Central Europe, Czech Republic) at an altitude of 950 m a.s.l. The aim of the experiment was to test the suitability of Alnus incana to form preparatory stands covering the site and thus enabling the reintroduction of more sensitive target species. A potential of Alnus incana to respond to slow-release fertilizing was tested as well. The control treatment showed sufficient growth dynamics, nevertheless, the fertilization significantly promoted the growth (documented by height, height increment and stem-base diameter). If some limitations of alder such as high light requirements are respected, the speckled alder can be recommended as a suitable species for preparatory stands even in the 7 th and 8 th altitudinal (vegetation) zones, especially when fertilized.

In experiments with willow planted in cadmium- or lead-contaminated Knopp nutrient solutions, the ability of willow and chelex gel in the diffusive gradient in a thin film (DGT) unit to absorb these elements was observed and compared. A method to compare the accumulated amount of metals by the willow and the sorbed amount of metal onto the DGT unit was designed. The accumulatedysorbed amount of metals was recalculated to the same units (m gc m � 2 day � 1 ). The study of Cd(Pb) dynamic uptake by willow showed an exponential decrease of metal content in the exposure solution. The willow planted in the Knopp nutrient solution accumulated Cd(Pb) less than did the DGT unit in the same matrix. The willow planted in the soilywater system accumulated more lead than did the willow in a lead-contaminated aquatic system nor did the DGT unit in the nitrate matrix. The willow further accumulated lead from the soilywater system due to the active co-uptake of nutrients and lead. The DGT technique was considered as very good comparative method to measure heavy metals plant uptake. This technique is suitable for use in water as well as in soil systems.

In the soil solutions obtained in situ with suction cups from soils (Cambisol and Fluvisol) of pot experiment with Salix smithiana Smith, Lolium perenne L. and Thlaspi caerulescens J. & C. Presl heavy metals species (Cd, Pb and Cu) were assayed by differential pulse anodic stripping voltammetry and diffusive gradient in thin films. Prediction of accumulation performed best at free metal ion concentrations in unchanged pH (in 10−3 mol L−1 NaClO4 base electrolyte). The speciation provided by differential pulse anodic stripping voltammetry according to pH can provide a detailed description of the soil solution matrix. The concentration of free metals in unchanged pH represents a small part of the total content and varied from 0.04 to 0.75% with two exceptions found for accumulating plants (the content of Cd2+ in the soil solution from T. caerulescens was about 6% and the content of Cu2+ in the soil solution from S. smithiana was about 30%). The available concentration as determined by diffusive gradient in thin films was not in correlation with the heavy metals concentration in plant biomass.

The article summarizes outcomes of a biomass study conducted in a young speckled alder plantation on a cold mountain site. At this location, the previously existing old forest was clear felled because of damage from air pollution, and present-day surface humus is in need of restoration. The intention of this study was to quantify the biomass and nutrients accumulated by alders and their components and assess whether the initial fertilization resulted in increased biomass production and nutrient accumulation in the biomass. Besides the control, two fertilized treatments were installed. In the surface treatment (SUT), the amendment was applied as a base dressing in small circles around trees. In the planting-hole treatment (PHT), the amendment was incorporated into soil inside the planting holes. Five growth seasons after planting and fertilization, six alders from each treatment were harvested including roots. Their biomass was quantified and analyzed for macroelements. The greatest pool of dry mass (DM) was branches in the control and stem wood in the fertilized treatments. The greatest pools of macroelements were leaves and branches. The most pronounced effects of fertilization were recorded in the DM and consequently in the absolute quantities of nutrients. The DM of an average tree in the control, SUT, and PHT was 85, 226, and 231 g, respectively. The absolute contents of nutrients per tree in the fertilized treatments showed the following increases, as compared with the control: (N) 2.5–2.6 times; (P) 1.6–2.4 times; (K) 1.8–2.1 times; and (Mg) 1.8–2.0 times, respectively. Speckled alder responded positively to fertilization.

led alder is a convenient pioneer species that can be cultivated on harsh mountain sites where the climatic extremes need to be alleviated and a layer of surface humus renewed. The initial slow-release fertilization is, nonetheless, highly desirable. As contrasted to the control the surface and planting hole applications of amendment reduced the total seven-year mortality rate by 9.5% and 20.1%, respectively, and the periodic annual height increment (2003-2009) was promoted by 47% and 59%, respectively. Analogous results were obtained when the values of basal stem diameter and crown diameter were compared. As for the method of application, the placement of the slow-release amendment in the planting holes seems to be more efficient mainly in terms of survival promoting. The surface application is, nonetheless, less laborious and also yielded satisfactory results.

The influence of N-S fertilizers on the copper content in the inter oilseed rape plants was studied in field expe riments. The evaluation involved two treatments of a single rate for the first spring fertilizer application with 100 kg N/ha in the AN treatment (nitrochalk) and 100 kg N/ha + 50 kg S/ha in the ANS treatment (ammonium nitrate and ammonium sulphate). A positive influence of the ANS fertilizer on the copper contents in different parts of plants was determined. The highest Cu concentrations were determined in the leaves and inflorescences, the lowest ones occurred in the stem. The concentration of Cu ranged within the interval of 1.56–8.75 mg Cu/kg of dry matter depending on the growth period and the part of the plant. No differences in copper content were determined in the seeds of individual treatment. The highest uptake in the above-ground parts of the plants was recorded in the green pod period and amounted to 57.4 g Cu/ha for the ANS treatment.