Investigation of a Perspective Urban Tree Species, Ginkgo biloba L., by Scientific Analysis of Historical Old Specimens.

In this study, we examined over 200-year-old Ginkgo biloba L. specimens under different environmental conditions. The overall aim was to explore which factors influence their vitality and general fitness in urban environments and thus their ability to tolerate stressful habitats. In order to determine this, we used a number of different methods, including histological examinations (stomatal density and size) and physiological measurements (peroxidase enzyme activity), as well as assessing the air pollution tolerance index (APTI). The investigation of the genetic relationships between individuals was performed using flow cytometry and miRNA marker methods. The genetic tests revealed that all individuals are diploid, whereas the lus-miR168 and lus-miR408 markers indicated a kinship relation between them. These results show that the effect of different habitat characteristics can be detected through morphological and physiological responses, thus indicating relatively higher stress values for all studied individuals. A significant correlation can be found between the level of adaptability and the relatedness of the examined individuals. These results suggest that Ginkgo biloba L. is well adapted to an environment with increased stress factors and therefore suitable for use in urban areas.

Light dependent differentiation of outdoors developed purple eggplant (Solanum melongena L.) pericarp layers: Leaf chlorenchyma characteristics of the pericarp layers dissected in the dark.

To interpret the final steps of chlorophyll biosynthesis, detailed knowledge of etiolation symptoms is necessary. Most of our knowledge originates from studies on plant materials grown in complete darkness. Hardly any information is available about the plastid development in internal parenchyma cells of fleshy fruits in which the food supply is almost unlimited. In this work, etiolation symptoms were studied in pericarp layers of purple eggplant (Solanum melongena L.). Tissue layers of fruits developed under open-air conditions and of etiolated fruits were dissected in a dark room. Transmission and 77 K fluorescence spectroscopy and ultrastructural studies were performed. Photosynthetic activities were measured and pigment contents were determined in light-grown fruits. The purple exocarp and a 1-1.5 cm wide green mesocarp layer of large fruits fully shade the internal pericarp layers, thus protochloropyll (ide) accumulated, flash-photoactive 644 and 655 nm emitting protochlorophyllide complexes, and only small amounts of chlorophylls were found. Photosynthetic activity was detected only in the external, green layer, which had fully developed chloroplasts, and showed 77 K fluorescence emission spectra characteristic for green leaves. The innermost endocarp regions and the etiolated fruits contained mainly protochlorophyll (ide), proplastids, and etioplasts, i.e. they showed etiolation symptoms. These symptoms correspond to those of leaves of dark-grown seedlings but are stable for long periods due to the almost unlimited nourishment supply from storage parenchyma cells. These results prove that the laboratory works with artificially dark-developed plant materials are good models of natural chlorophyll biosynthesis and plastid development.

Revealing the nuclearity of iron citrate complexes at biologically relevant conditions.

Citric acid plays an ubiquitous role in the complexation of essential metals like iron and thus it has a key function making them biologically available. For this, iron(III) citrate complexes are considered among the most significant coordinated forms of ferric iron that take place in biochemical processes of all living organisms. Although these systems hold great biological relevance, their coordination chemistry has not been fully elucidated yet. The current study aimed to investigate the speciation of iron(III) citrate using Mössbauer and electron paramagnetic resonance spectroscopies. Our aim was to gain insights into the structure and nuclearity of the complexes depending on the pH and iron to citrate ratio. By applying the frozen solution technique, the results obtained directly reflect the iron speciation present in the aqueous solution. At 1:1 iron:citrate molar ratio, polynuclear species prevailed forming most probably a trinuclear structure. In the case of citrate excess, the coexistence of several monoiron species with different coordination environments was confirmed. The stability of the polynuclear complexes was checked in the presence of organic solvents.

Iron uptake of etioplasts is independent from photosynthesis but applies the reduction-based strategy.

Introduction: Iron (Fe) is one of themost important cofactors in the photosynthetic apparatus, and its uptake by chloroplasts has also been associated with the operation of the photosynthetic electron transport chain during reduction-based plastidial Fe uptake. Therefore, plastidial Fe uptake was considered not to be operational in the absence of the photosynthetic activity. Nevertheless, Fe is also required for enzymatic functions unrelated to photosynthesis, highlighting the importance of Fe acquisition by non-photosynthetic plastids. Yet, it remains unclear how these plastids acquire Fe in the absence of photosynthetic function. Furthermore, plastids of etiolated tissues should already possess the ability to acquire Fe, since the biosynthesis of thylakoid membrane complexes requires a massive amount of readily available Fe. Thus, we aimed to investigate whether the reduction-based plastidial Fe uptake solely relies on the functioning photosynthetic apparatus. Methods: In our combined structure, iron content and transcript amount analysis studies, we used Savoy cabbage plant as a model, which develops natural etiolation in the inner leaves of the heads due to the shading of the outer leaf layers. Results: Foliar and plastidial Fe content of Savoy cabbage leaves decreased towards the inner leaf layers. The leaves of the innermost leaf layers proved to be etiolated, containing etioplasts that lacked the photosynthetic machinery and thus were photosynthetically inactive. However, we discovered that these etioplasts contained, and were able to take up, Fe. Although the relative transcript abundance of genes associated with plastidial Fe uptake and homeostasis decreased towards the inner leaf layers, both ferric chelate reductase FRO7 transcripts and activity were detected in the innermost leaf layer. Additionally, a significant NADP(H) pool and NAD(P)H dehydrogenase activity was detected in the etioplasts of the innermost leaf layer, indicating the presence of the reducing capacity that likely supports the reduction-based Fe uptake of etioplasts. Discussion: Based on these findings, the reduction-based plastidial Fe acquisition should not be considered exclusively dependent on the photosynthetic functions.

Antioxidative Defense, Suppressed Nitric Oxide Accumulation, and Synthesis of Protective Proteins in Roots and Leaves Contribute to the Desiccation Tolerance of the Resurrection Plant Haberlea rhodopensis.

The desiccation tolerance of plants relies on defense mechanisms that enable the protection of macromolecules, biological structures, and metabolism. Although the defense of leaf tissues exposed to solar irradiation is challenging, mechanisms that protect the viability of the roots, yet largely unexplored, are equally important for survival. Although the photosynthetic apparatus in leaves contributes to the generation of oxidative stress under drought stress, we hypothesized that oxidative stress and thus antioxidative defense is also predominant in the roots. Thus, we aimed for a comparative analysis of the protective mechanisms in leaves and roots during the desiccation of Haberlea rhodopensis. Consequently, a high content of non-enzymatic antioxidants and high activity of antioxidant enzymes together with the activation of specific isoenzymes were found in both leaves and roots during the final stages of desiccation of H. rhodopensis. Among others, catalase and glutathione reductase activity showed a similar tendency of changes in roots and leaves, whereas, unlike that in the leaves, superoxide dismutase activity was enhanced under severe but not under medium desiccation in roots. Nitric oxide accumulation in the root tips was found to be sensitive to water restriction but suppressed under severe desiccation. In addition to the antioxidative defense, desiccation induced an enhanced abundance of dehydrins, ELIPs, and sHSP 17.7 in leaves, but this was significantly better in roots. In contrast to leaf cells, starch remained in the cells of the central cylinder of desiccated roots. Taken together, protective compounds and antioxidative defense mechanisms are equally important in protecting the roots to survive desiccation. Since drought-induced damage to the root system fundamentally affects the survival of plants, a better understanding of root desiccation tolerance mechanisms is essential to compensate for the challenges of prolonged dry periods.

Histological and Physiological Effects of Treatment of Rudbeckia hirta with Gamma Radiation.

The breeding of resistant, high-yield, decorative ornamental plant varieties may be impacted by climate change in the future. The use of radiation induces mutations in plants, thereby increasing the genetic variability of plant species. Rudbeckia hirta has long been a very popular species in urban green space management. The goal is to examine whether gamma mutation breeding can be applied to the breeding stock. Specifically, differences were measured between the M1 and M2 generations, as well as the effect of different radiation doses belonging to the same generation. Morphological measurements showed that gamma radiation has an effect on the measured parameters in several cases (larger crop size, faster development, larger number of trichomes). Physiological measurements (examination of chlorophyll and carotenoid content, POD activity, and APTI) also showed a beneficial effect of radiation, especially at higher doses (30 Gy), for both tested generations. The treatment was also effective in the case of 45 Gy, but this radiation dose resulted in lower physiological data. The measurements show that gamma radiation has an effect on the Rudbeckia hirta strain and may play a role in breeding in the future.

Functional Analysis of Chloroplast Iron Uptake and Homeostasis.

Iron has a crucial role in plastid biology. Iron is a required cofactor for the operation of the photosynthetic functions and other metabolic pathways. Despite the importance of the iron homeostasis in chloroplasts, the functional analysis of the plastidial iron uptake and homeostasis still lack a consensus methodology. Here, we describe a sequence of subsequent techniques that can be applied in functional characterization of proteins involved in iron uptake and incorporation into chloroplasts as well as of the non-transport protein members of the chloroplast iron homeostasis. Since the ferrous iron ligation of bathophenantroline disulfonate is specific and not disrupted by the presence of other transition metals, it offers a simple way for iron quantification both in solubilized chloroplast samples as well as in ferric chelate reductase activity measurements.

Protein Changes in Shade and Sun Haberlea rhodopensis Leaves during Dehydration at Optimal and Low Temperatures.

Haberlea rhodopensis is a unique resurrection plant of high phenotypic plasticity, colonizing both shady habitats and sun-exposed rock clefts. H. rhodopensis also survives freezing winter temperatures in temperate climates. Although survival in conditions of desiccation and survival in conditions of frost share high morphological and physiological similarities, proteomic changes lying behind these mechanisms are hardly studied. Thus, we aimed to reveal ecotype-level and temperature-dependent variations in the protective mechanisms by applying both targeted and untargeted proteomic approaches. Drought-induced desiccation enhanced superoxide dismutase (SOD) activity, but FeSOD and Cu/ZnSOD-III were significantly better triggered in sun plants. Desiccation resulted in the accumulation of enzymes involved in carbohydrate/phenylpropanoid metabolism (enolase, triosephosphate isomerase, UDP-D-apiose/UDP-D-xylose synthase 2, 81E8-like cytochrome P450 monooxygenase) and protective proteins such as vicinal oxygen chelate metalloenzyme superfamily and early light-induced proteins, dehydrins, and small heat shock proteins, the latter two typically being found in the latest phases of dehydration and being more pronounced in sun plants. Although low temperature and drought stress-induced desiccation trigger similar responses, the natural variation of these responses in shade and sun plants calls for attention to the pre-conditioning/priming effects that have high importance both in the desiccation responses and successful stress recovery.

Apoplast utilisation of nanohaematite initiates parallel suppression of RIBA1 and FRO1&3 in Cucumis sativus.

Nanoscale Fe containing particles can penetrate the root apoplast. Nevertheless, cell wall size exclusion questions that for Fe mobilisation, a close contact between the membrane integrating FERRIC REDUCTASE OXIDASE (FRO) enzymes and Fe containing particles is required. Haematite nanoparticle suspension, size of 10-20 nm, characterized by 57Fe Mössbauer spectroscopy, TEM, ICP and SAED was subjected to Fe utilisation by the flavin secreting model plant cucumber (Cucumis sativus). Alterations in the structure and distribution of the particles were revealed by 57Fe Mössbauer spectroscopy, HRTEM and EDS element mapping. Biological utilisation of Fe resulted in a suppression of Fe deficiency responses (expression of CsFRO 1, 2 & 3 and RIBOFLAVIN A1; CsRIBA1 genes and root ferric chelate reductase activity). Haematite nanoparticles were stacked in the middle lamella of the apoplast. Fe mobilisation is evidenced by the reduction in the particle size. Fe release from nanoparticles does not require a contact with the plasma membrane. Parallel suppression in the CsFRO 1&3 and CsRIBA1 transcript amounts support that flavin biosynthesis is an inclusive Fe deficiency response involved in the reduction-based Fe utilisation of Cucumis sativus roots. CsFRO2 is suggested to play a role in the intracellular Fe homeostasis.

Iron Status Affects the Zinc Accumulation in the Biomass Plant Szarvasi-1.

Thinopyrum obtusiflorum (syn. Elymus elongatus subsp. ponticus) cv. Szarvasi-1 (Poaceae, Triticeae) is a biomass plant with significant tolerance to certain metals. To reveal its accumulation capacity, we investigated its Zn uptake and tolerance in a wide range: 0.2 to 1000 µM Zn concentration. The root and shoot weight, shoot length, shoot water content and stomatal conductance proved to be only sensitive to the highest applied Zn concentrations, whereas the concentration of malondialdehyde increased only at the application of 1 mM Zn in the leaves. Although physiological status proved to be hardy against Zn exposure, shoot Zn content significantly increased in parallel with the applied Zn treatment, reaching the highest Zn concentration at 1.9 mg g-1 dry weight. The concentration of K, Mg and P considerably decreased in the shoot at the highest Zn exposures, where that of K and P also correlated with a decrease in water content. Although the majority of microelements remained unaffected, Mn decreased in the root and Fe content had a negative correlation with Zn both in the shoot and root. In turn, the application of excessive EDTA maintained a proper Fe supply for the plants but lowered Zn accumulation both in roots and shoots. Thus, the Fe-Zn competition for Fe chelating phytosiderophores and/or for root uptake transporters fundamentally affects the Zn accumulation properties of Szarvasi-1. Indeed, the considerable Zn tolerance of Szarvasi-1 has a high potential in Zn accumulation.

S-Methylmethionine Effectively Alleviates Stress in Szarvasi-1 Energy Grass by Reducing Root-to-Shoot Cadmium Translocation.

S-methylmethionine (SMM) is a universal metabolite of higher plants derived from L-methionine that has an approved priming effect under different types of abiotic and biotic stresses. Szarvasi-1 energy grass (Elymus elongatus subsp. ponticus cv. Szarvasi-1) is a biomass plant increasingly applied in phytoremediation to stabilize or extract heavy metals. In this study, Szarvasi-1 was grown in a nutrient solution. As a priming agent, SMM was applied in 0.02, 0.05 and 0.1 mM concentrations prior to 0.01 mM Cd addition. The growth and physiological parameters, as well as the accumulation pattern of Cd and essential mineral nutrients, were investigated. Cd exposure decreased the root and shoot growth, chlorophyll concentration, stomatal conductance, photosystem II function and increased the carotenoid content. Except for stomatal conductance, SMM priming had a positive effect on these parameters compared to Cd treatment without priming. In addition, it decreased the translocation and accumulation of Cd. Cd treatment decreased K, Mg, Mn, Zn and P in the roots, and K, S, Cu and Zn in the shoots compared to the untreated control. SMM priming changed the pattern of nutrient uptake, of which Fe showed characteristic accumulation in the roots in response to increasing SMM concentrations. We have concluded that SMM priming exerts a positive effect on Cd-stressed Szarvasi-1 plants, which retained their physiological performance and growth. This ameliorative effect is suggested to be based on, at least partly, the lower root-to-shoot Cd translocation by the upregulated Fe uptake and transport.

Reactivation of the Photosynthetic Apparatus of Resurrection Plant Haberlea rhodopensis during the Early Phase of Recovery from Drought- and Freezing-Induced Desiccation.

Haberlea rhodopensis is a unique desiccation-tolerant angiosperm that also survives winter frost. As, upon freezing temperatures, H. rhodopensis desiccates, the taxon is proposed to survive low temperature stress using its desiccation tolerance mechanisms. To reveal the validity of this hypothesis, we analyzed the structural alterations and organization of photosynthetic apparatus during the first hours of recovery after drought- and freezing-induced desiccation. The dynamics of the ultrastructure remodeling in the mesophyll cells and the restoration of the thylakoid membranes shared similarities independent of the reason for desiccation. Among the most obvious changes in thylakoid complexes, the proportion of the PSI-LHCII complex strongly increased around 70% relative water content (RWC), whereas the proportion of Lhc monomers decreased from the beginning of rehydration. We identified enhanced levels of cyt b6f complex proteins that contributed to the enhanced electron flow. The high abundance of proteins related to excitation energy dissipation, PsbS, Lhcb5, Lhcb6 and ELIPs, together with the increased content of dehydrins contributed to the preservation of cellular integrity. ELIP expression was maintained at high levels up to 9 h into recovery. Although the recovery processes from drought- and freezing-induced desiccation were found to be similar in progress and time scale, slight variations indicate that they are not identical.

Qualitative and quantitative evaluation of thylakoid complexes separated by Blue Native PAGE.

BACKGROUND: Blue Native polyacrylamide gel electrophoresis (BN PAGE) followed by denaturing PAGE is a widely used, convenient and time efficient method to separate thylakoid complexes and study their composition, abundance, and interactions. Previous analyses unravelled multiple monomeric and dimeric/oligomeric thylakoid complexes but, in certain cases, the separation of complexes was not proper. Particularly, the resolution of super- and megacomplexes, which provides important information on functional interactions, still remained challenging. RESULTS: Using a detergent mixture of 1% (w/V) n-dodecyl-ß-D-maltoside plus 1% (w/V) digitonin for solubilisation and 4.3-8% gel gradients for separation as methodological improvements in BN PAGE, several large photosystem (PS) I containing bands were detected. According to BN(/BN)/SDS PAGE and mass spectrometry analyses, these PSI bands proved to be PSI-NADH dehydrogenase-like megacomplexes more discernible in maize bundle sheath thylakoids, and PSI complexes with different light-harvesting complex (LHC) complements (PSI-LHCII, PSI-LHCII*) more abundant in mesophyll thylakoids of lincomycin treated maize. For quantitative determination of the complexes and their comparison across taxa and physiological conditions, sample volumes applicable to the gel, correct baseline determination of the densitograms, evaluation methods to resolve complexes running together, calculation of their absolute/relative amounts and distribution among their different forms are proposed. CONCLUSIONS: Here we report our experience in Blue/Clear-Native polyacrylamide gel electrophoretic separation of thylakoid complexes, their identification, quantitative determination and comparison in different samples. The applied conditions represent a powerful methodology for the analysis of thylakoid mega- and supercomplexes.

New aspects of the photodegradation of iron(III) citrate: spectroscopic studies and plant-related factors.

Iron (Fe) is an essential cofactor for all livings. Although Fe membrane transport mechanisms often utilize FeII, uncoordinated or deliberated ferrous ions can initiate Fenton reactions. FeIII citrate complexes are among the most important complexed forms of FeIII especially in plants that, indeed, can undergo photoreduction. Since leaves as photosynthetic organs of higher plants are generally exposed to illumination in daytime, photoreaction of ferric species may have biological relevance in iron metabolism, the relevance of which is poorly understood. In present work FeIII citrate transformation during the photodegradation in solution and after foliar application on leaves was studied by Mössbauer analysis directly. To obtain irradiation time dependence of the speciation of iron in solutions, four model solutions of different pH values (1.5, 3.3, 5.5, and 7.0) with Fe to citrate molar ratio 1:1.1 were exposed to light. Highly acidic conditions led to a complete reduction of Fe together with the formation of FeII citrate and hexaaqua complexes in equal concentration. At higher pH, the only product of the photodegradation was FeII citrate, which was later reoxidized and polymerized, resulting in the formation of polynuclear stable ferric compound. To test biological relevance, leaves of cabbage were treated with FeIII citrate solution. X-ray fluorescence imaging indicated the accumulation of Fe in the treated leaf parts. Mössbauer analysis revealed the presence of several ferric species incorporated into the biological structure. The Fe speciation observed should be considered in biological systems where FeIII citrate has a ubiquitous role in Fe acquisition and homeostasis.

Iron in leaves: chemical forms, signalling, and in-cell distribution.

Iron (Fe) is an essential transition metal. Based on its redox-active nature under biological conditions, various Fe compounds serve as cofactors in redox enzymes. In plants, the photosynthetic machinery has the highest demand for Fe. In consequence, the delivery and incorporation of Fe into cofactors of the photosynthetic apparatus is the focus of Fe metabolism in leaves. Disturbance of foliar Fe homeostasis leads to impaired biosynthesis of chlorophylls and composition of the photosynthetic machinery. Nevertheless, mitochondrial function also has a significant demand for Fe. The proper incorporation of Fe into proteins and cofactors as well as a balanced intracellular Fe status in leaf cells require the ability to sense Fe, but may also rely on indirect signals that report on the physiological processes connected to Fe homeostasis. Although multiple pieces of information have been gained on Fe signalling in roots, the regulation of Fe status in leaves has not yet been clarified in detail. In this review, we give an overview on current knowledge of foliar Fe homeostasis, from the chemical forms to the allocation and sensing of Fe in leaves.

Supraoptimal Iron Nutrition of Brassica napus Plants Suppresses the Iron Uptake of Chloroplasts by Down-Regulating Chloroplast Ferric Chelate Reductase.

Iron (Fe) is an essential micronutrient for plants. Due to the requirement for Fe of the photosynthetic apparatus, the majority of shoot Fe content is localised in the chloroplasts of mesophyll cells. The reduction-based mechanism has prime importance in the Fe uptake of chloroplasts operated by Ferric Reductase Oxidase 7 (FRO7) in the inner chloroplast envelope membrane. Orthologue of Arabidopsis thaliana FRO7 was identified in the Brassica napus genome. GFP-tagged construct of BnFRO7 showed integration to the chloroplast. The time-scale expression pattern of BnFRO7 was studied under three different conditions: deficient, optimal, and supraoptimal Fe nutrition in both leaves developed before and during the treatments. Although Fe deficiency has not increased BnFRO7 expression, the slight overload in the Fe nutrition of the plants induced significant alterations in both the pattern and extent of its expression leading to the transcript level suppression. The Fe uptake of isolated chloroplasts decreased under both Fe deficiency and supraoptimal Fe nutrition. Since the enzymatic characteristics of the ferric chelate reductase (FCR) activity of purified chloroplast inner envelope membranes showed a significant loss for the substrate affinity with an unchanged saturation rate, protein level regulation mechanisms are suggested to be also involved in the suppression of the reduction-based Fe uptake of chloroplasts together with the saturation of the requirement for Fe.

The developmental and iron nutritional pattern of PIC1 and NiCo does not support their interdependent and exclusive collaboration in chloroplast iron transport in Brassica napus.

MAIN CONCLUSION: The accumulation of NiCo following the termination of the accumulation of iron in chloroplast suggests that NiCo is not solely involved in iron uptake processes of chloroplasts. Chloroplast iron (Fe) uptake is thought to be operated by a complex containing permease in chloroplast 1 (PIC1) and nickel-cobalt transporter (NiCo) proteins, whereas the role of other Fe homeostasis-related transporters such as multiple antibiotic resistance protein 1 (MAR1) is less characterized. Although pieces of information exist on the regulation of chloroplast Fe uptake, including the effect of plant Fe homeostasis, the whole system has not been revealed in detail yet. Thus, we aimed to follow leaf development-scale changes in the chloroplast Fe uptake components PIC1, NiCo and MAR1 under deficient, optimal and supraoptimal Fe nutrition using Brassica napus as model. Fe deficiency decreased both the photosynthetic activity and the Fe content of plastids. Supraoptimal Fe nutrition caused neither Fe accumulation in chloroplasts nor any toxic effects, thus only fully saturated the need for Fe in the leaves. In parallel with the increasing Fe supply of plants and ageing of the leaves, the expression of BnPIC1 was tendentiously repressed. Though transcript and protein amount of BnNiCo tendentiously increased during leaf development, it was even markedly upregulated in ageing leaves. The relative transcript amount of BnMAR1 increased mainly in ageing leaves facing Fe deficiency. Taken together chloroplast physiology, Fe content and transcript amount data, the exclusive participation of NiCo in the chloroplast Fe uptake is not supported. Saturation of the Fe requirement of chloroplasts seems to be linked to the delay of decomposing the photosynthetic apparatus and keeping chloroplast Fe homeostasis in a rather constant status together with a supressed Fe uptake machinery.

Chloroplasts preferentially take up ferric-citrate over iron-nicotianamine complexes in Brassica napus.

MAIN CONCLUSION: Fe uptake machinery of chloroplasts prefers to utilise Fe(III)-citrate over Fe-nicotianamine complexes. Iron uptake in chloroplasts is a process of prime importance. Although a few members of their iron transport machinery were identified, the substrate preference of the system is still unknown. Intact chloroplasts of oilseed rape (Brassica napus) were purified and subjected to iron uptake studies using natural and artificial iron complexes. Fe-nicotianamine (NA) complexes were characterised by 5 K, 5 T Mössbauer spectrometry. Expression of components of the chloroplast Fe uptake machinery was also studied. Fe(III)-NA contained a minor paramagnetic Fe(II) component (ca. 9%), a paramagnetic Fe(III) component exhibiting dimeric or oligomeric structure (ca. 20%), and a Fe(III) complex, likely being a monomeric structure, which undergoes slow electronic relaxation at 5 K (ca. 61%). Fe(II)-NA contained more than one similar chemical Fe(II) environment with no sign of Fe(III) components. Chloroplasts preferred Fe(III)-citrate compared to Fe(III)-NA and Fe(II)-NA, but also to Fe(III)-EDTA and Fe(III)-o,o'EDDHA, and the Km value was lower for Fe(III)-citrate than for the Fe-NA complexes. Only the uptake of Fe(III)-citrate was light-dependent. Regarding the components of the chloroplast Fe uptake system, only genes of the reduction-based Fe uptake system showed high expression. Chloroplasts more effectively utilize Fe(III)-citrate, but hardly Fe-NA complexes in Fe uptake.

Response of soybean plants to the application of synthetic and biodegradable Fe chelates and Fe complexes.

The growing concern over the environmental risk of synthetic chelate application promotes the search for alternatives in Fe fertilization, such as biodegradable chelating agents and natural complexing agents. In this work, plant responses to the application of several Fe treatments (chelates and complexes) was analyzed to study their potential use in Fe fertilization under calcareous conditions. Thus, the root ferric chelate reductase (FCR) activity of soybean (Glycine max cv. Klaxon) plants was determined, and the effectiveness of the Fe chelates and complexes assessed in a pot experiment, by SPAD and fluorescence induction measurements, and the determination of Fe distribution in plant and soil. Additionally, 57Fe Mössbauer spectroscopy was conducted to identify the Fe forms present in the soybean roots. The highest FCR activity was observed for the chelates EDDS/Fe3+ and IDHA/Fe3+; while no activity was observed when using complexes as Fe substrates. In contrast to the FCR data, the pot experiment confirmed that the o,oEDDHA/Fe3+ is the most effective treatment, and the complexes LS/Fe3+ and GA/Fe3+ are able to alleviate Fe chlorosis, also indicated by SPAD data and the maximal quantum efficiency of photosystem II reaction centers as vitality parameters, and the enhanced plant uptake of Fe from natural sources.