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Anthropogenic pollution poses a major threat to aquatic ecosystems, which can lead to their degradation. The accumulation and toxicity of metals and trace elements in fish leads to physiological and chemical changes in the fish body. In this study, we investigated the effects of bioaccumulation of metals and trace elements on freshwater fish fatty acid profiles at two different sites before and after the discharge of untreated municipal wastewater in two fish species with different diet habits. Although the concentrations of toxic elements were below the maximum levels proposed by the EU and the Republic of Serbia, this study showed statistically significant correlations between the presence of certain elements and the fatty acid (FA) profile in fish muscle. Lower concentrations of polyunsaturated FA in fish sampled after the discharge of untreated municipal wastewater were detected., In this study, the concentrations of metals and trace elements (As, Cd, Co, Cr, Cu, Hg, Ni, Pb and Zn) were determined in the muscle tissue of adult roach and white bream at two different sites in the Belgrade section of the Danube. Twenty-six fatty acids, consisting of nine saturated FA (SFAs), seven monosaturated FA (MUFAs) and ten polysaturated FA (PUFAs), were identified. The analysis of the concentration of metals and trace elements of the roach and white bream showed species-specific differences in their bioaccumulation. Four of all elements analyzed (As, Hg, Ni and Pb) correlated significantly with the changes in FA profiles in fish from both sampling sites, with the exception of Cu, which correlated with the FA profile at the site before, and Zn, whose concentration influenced the FA profile at the site after wastewater discharges. The lower PUFA content in the fish from a site under higher environment pressure could indicate that the fish are stressed. The results suggest that changes in lipid composition may be one of the protective mechanisms of cells to cope with anthropogenic stressors.

Haematococcus pluvialis is a unicellular green alga with a complex life cycle and a remarkable metabolic and structural adaptability that allows it to thrive in metal-infested environments. H. pluvialis could be potentially used in the remediation of waters polluted with metals, such as manganese (Mn). Mn is also an essential element involved in different metabolic processes, such as photosynthesis and antioxidantive defence. Herein, we examined morphological response of metabolically active green cell type of H. pluvialis (microzooids) to high Mn concentrations exceeding their physiological quota. When exposed to 1 mM Mn2+, cell viability remained stable over a 3-day period. Inductively coupled plasma atomic emission spectroscopy showed the prompt uptake of Mn by the microzooid cells after 1 h of the treatment, with a modest increase of the concentration of Mn in the biomass at 24 h. Scanning electron microscopy revealed granular deposits on microzooid surfaces after 1 hour, likely Mn deposits, while transmission electron microscopy (TEM) micrographs showed that some cells had wall rupture and degraded intracellular content and damaged organelles. After 24 and 72 h, a different type of cell morphology emerged, characterized by thickened cell wall, preserved intracellular compartments, and reduced total area of lipid droplets. Both cell types exhibited vacuoles containing dark granules, possibly indicative of Mn accumulations. Quantitative TEM analysis demonstrated that an excess of Mn reduced cell cross-section and lipid droplet area while increasing vacuole cross-section and cell wall thickness. The intricate adaptive responses of H. pluvialis to elevated Mn concentrations exemplified by cell wall thickening, reduction in lipid droplets total area due to increased energy demand, and the accumulation of Mn in vacuoles, exhibits the impressive structural adaptability. Further investigation using analytical methods will provide a more profound understanding of the me

Heavy metals remain a major pollutant in waters near mining sites. Water pollution is a current, longterm problem that affects plants and organisms that live in these water systems, and the effect is very harmful not only for individual species and populations, but also for the entire biological community. This study analyzed the potential of the microalga Chlorella sorokiniana in the adsorption of four selected metals: Fe, Cu, Zn and Ni, all present in high amounts in Lake Robule. This pilot study was conducted to evaluate the potential of these microalgae for possible use in future bioremediation treatment of these water. A laboratory study of metal accumulation in lake water samples lasted for 7 days, after which a total decrease in metal concentration was observed, namely Fe ~25%, Cu ~17%, and Zn ~4% on the seventh day. This study confirms the strong potential of microalgae C. sorokiniana to reduce the presence of heavy metals in conditions known for the low pH value of water with a high percentage of Fe and other heavy metals.

Metabolism of metals in microalgae and adaptation to metal excess are of significant environmental importance. We report a three-step mechanism that the green microalga Chlorella sorokiniana activates during the acquisition of and adaptation to manganese (Mn), which is both an essential trace metal and a pollutant of waters. In the early stage, Mn2+ was mainly bound to membrane phospholipids and phosphates in released mucilage. The outer cell wall was reorganized and lipids were accumulated, with a relative increase in lipid saturation. Intracellular redox settings were rapidly altered in the presence of Mn excess, with increased production of reactive oxygen species that resulted in lipid peroxidation and a decrease in the concentration of thiols. In the later stage, Mn2+ was chelated by polyphosphates and accumulated in the cells. The structure of the inner cell wall was modified and the redox milieu established a new balance. Polyphosphates serve as a transient Mn2+ storage ligand, as proposed previously. In the final stage, Mn was stored in multivalent Mn clusters that resemble the structure of the tetramanganese–calcium core of the oxygen-evolving complex. The present findings elucidate the bioinorganic chemistry and metabolism of Mn in microalgae, and may shed new light on water-splitting Mn clusters.

Metabolism of metals in microalgae, as well as their adaptation to metal excess, are of significant environmental importance. We have found previously that the green microalga Chlorella sorokiniana accumulates environmental Mn excess in the form of a multivalent Mn-O-Ca cluster with structure that is very similar of oxygen-evolving complex (OEC) in photosystem II1. The application of microalgae in the ‘green’ synthesis of catalytic metal clusters is very important, since the use of toxic chemicals for traditional synthesis could be avoided. The aim of this study was to investigate the magnetic, redox and structural properties of this cluster. The magnetic properties of the cluster, were tested using Low-T-EPR spectroscopy and SQUID magnetometry. Based on the analysis of the EPR spectra, it was concluded that the spin of Mn in the cluster is >1/2. In addition, the spin of Mn is not an integer since parallel mode EPR did not deliver any detectable spectrum. The paramagnetic nature of the Mn-O-Ca cluster was confirmed by the SQUID instrument. Effective magnetic moment calculated per ion Mn was ~5 BM (Mn ratio in biomass was 12,1%). For the investigation of redox and structural properties, an extract of cluster from microalga was used. Cluster extraction involved the application of a series of solvents - phenol in Tris-Cl buffer, chloroform, methanol, 0,17% sodium hypochlorite2. Cyclic and differential voltagrams were recorded in the range from -1,5 to 2 mV3. The analysis revealed the presence of different oxidation states of Mn (+2, +3, +4). The peak potentials resembled the potentials in OEC model compounds2.The redox similarities between the Mn-O-Ca cluster in microalgae and OEC imply that cluster may be have a similar catalytic activity. More detailed analysis of activity and structure of Mn-O-Ca cluster is warranted.

Nickel is utilized by microalgae as a co-factor of urease. On the other hand, this transition metal represents an important pollutant of aquatic ecosystems. The effects of Ni(II) excess on microalgae and the mechanisms of adaptation are poorly understood. Redox processes represent an important component of the mechanisms of interaction of microalgae with transition metals. Pertinent to this, we analyzed redox changes in Chlorella sorokiniana culture that are induced by high levels of Ni(II). The intracellular level of reactive oxygen species (ROS) showed a rapid two-phase increase that took place prior to Ni accumulation in the cell. This was accompanied by oxidation of thiols and drastic deglutathyonilation of proteins. PAM fluorimetry showed that Ni excess induced an increase in the efficiency of photosystem II and promoted electron flow in chloroplasts, which is most likely responsible for ROS rise. In addition, a rising trend in the chlorophyll level was observed. On the other hand, the level of lipid peroxidation and activities of key antioxidative enzymes were not increased, which implies that oxidative stress is not an important player in Ni adaptation/toxicity. After prolonged exposure the efficiency of photosystem II drops, nickel is accumulated in the cells, and new redox balance is established. Our results imply that redox signalling is involved in Ni-induced metabolic activation and that key changes take place in photosynthetic machinery.

METHODS The impact of a set of concentrations of Mn2+ and Ni2+ on growth of C. sorokiniana culture in 3N-BBM+V medium in the early stationary phase was evaluated by changes in optical density at 750 nm and biomass during 7 days treatment. Mucilage release was analyzed using SEM microscopy. Redox settings were analyzed by oxidation-sensitive fluorescent probe and assays for thiols. RESULTS Ni was more toxic than Mn and affected culture growth at lower concentrations. Microalgal cells started releasing mucilage polymers within 1 h of exposure to 1 mM Mn2+, whereas no mucilage was observed even at 24 h of treatment with equimolar Ni2+. The peak of reactive oxygen species production was reached faster for Ni2+ than Mn2+. Mn-induced drops in the concentration of reduced thiols showed a recovery after 1 h and 24 h. Ni2+-induced drop was irreversible. The observed differences between the impact of Mn and Ni may be related to different redox and coordinative properties and to higher capacities of microalgae to sequester Mn in relation to higher quotas than Ni that are required for normal functio

There is a significant interest in understanding coordination and photo-chemistry of tetracycline antibiotics, primarily in relation to the development of advanced oxidation processes for degradation of these pollutants in water processing. Herein we analyzed the pH-dependence of interactions of tetracycline with ferric iron and photosensitivity of tetracycline to UV-A and UV-B, using a set of methods – UV–vis, Raman and electron paramagnetic resonance spectroscopy, MS spectrometry, HPLC, and cyclic voltammetry. Tetracycline and Fe3+ mainly bind through amide and OH groups in tricarbonylamide moiety to form a stable complex with 1:1 stoichiometry at pH ≤ 5. The interaction is reversible and tetracycline is released from the complex with pH increase. Tetracycline in the complex is stabilized and less susceptible than free tetracycline to oxidation by hydroxyl radical that is produced by UV-induced photolysis of Fe3+–OH- complexes. Redox properties of tetracycline were altered with increasing pH and it showed increased susceptibility to UV-induced degradation. In close, the system composed of tetracycline and ferric iron shows coordination and photo-redox chemistry that is dependent of pH in relation to the solubility of Fe3+ species and protonation of tetracycline. The development and optimization of advanced oxidation processes should take into account that iron may bind and stabilize pollutants and that the redox landscape of water changes drastically with pH.

The effects of a fructose-rich diet and chronic stress on copper metabolism in the kidneys are still understudied. We investigated whether fructose and/or chronic unpredictable stress modulate copper metabolism in a way that affects redox homeostasis, thus contributing to progression of metabolic disturbances in the kidney. We determined protein level of copper transporters, chaperones, and cuproenzymes including cytochrome c oxidase, as well as antioxidant enzymes function in the kidneys of male Wistar rats subjected to 20% liquid fructose supplementation and/or chronic stress. Liquid fructose supplementation increased level of copper chaperone of superoxide dismutase and decreased metallothionein level, while rendering the level of copper importer and copper chaperones involved in copper delivery to mitochondria and trans Golgi network unaffected. Stress had no effect on renal copper metabolism. The activity and expression of renal antioxidant enzymes remained unaltered in all experimental groups. In conclusion, fructose, independently of stress, decreased renal copper level, and modulated renal copper metabolism as to preserve vital cellular function including mitochondrial energy production and antioxidative defense, at the expense of intracellular copper storage.

In this contribution, four new compounds synthesized from 4-hydroxycoumarin and tyramine/octopamine/norepinephrine/3-methoxytyramine are characterized spectroscopically (IR and NMR), chromatographically (UHPLC-DAD), and structurally at the B3LYP/6-311++G*(d,p) level of theory. The crystal structure of the 4-hydroxycoumarin-octopamine derivative was solved and used as a starting geometry for structural optimization. Along with the previously obtained 4-hydroxycoumarin-dopamine derivative, the intramolecular interactions governing the stability of these compounds were quantified by NBO and QTAIM analyses. Condensed Fukui functions and the HOMO-LUMO gap were calculated and correlated with the number and position of OH groups in the structures. In vitro cytotoxicity experiments were performed to elucidate the possible antitumor activity of the tested substances. For this purpose, four cell lines were selected, namely human colon cancer (HCT-116), human adenocarcinoma (HeLa), human breast cancer (MDA-MB-231), and healthy human lung fibroblast (MRC-5) lines. A significant selectivity towards colorectal carcinoma cells was observed. Molecular docking and molecular dynamics studies with carbonic anhydrase, a prognostic factor in several cancers, complemented the experimental results. The calculated MD binding energies coincided well with the experimental activity, and indicated 4-hydroxycoumarin-dopamine and 4-hydroxycoumarin-3-methoxytyramine as the most active compounds. The ecotoxicology assessment proved that the obtained compounds have a low impact on the daphnia, fish, and green algae population.

Two Balkan Peninsula endemics, Nepeta rtanjensis and N. argolica subsp. argolica, both characterized by specialized metabolite profiles predominated by iridoids and phenolics, are differentiated according to the stereochemistry of major iridoid aglycone nepetalactone (NL). For the first time, the present study provides a comparative analysis of antimicrobial and immunomodulating activities of the two Nepeta species and their major iridoids isolated from natural sources—cis,trans-NL, trans,cis-NL, and 1,5,9-epideoxyloganic acid (1,5,9-eDLA), as well as of phenolic acid rosmarinic acid (RA). Methanol extracts and pure iridoids displayed excellent antimicrobial activity against eight strains of bacteria and seven strains of fungi. They were especially potent against food-borne pathogens such as L. monocytogenes, E. coli, S. aureus, Penicillium sp., and Aspergillus sp. Targeted iridoids were efficient agents in preventing biofilm formation of resistant P. aeruginosa strain, and they displayed additive antimicrobial interaction. Iridoids are, to a great extent, responsible for the prominent antimicrobial activities of the two Nepeta species, although are probably minor contributors to the moderate immunomodulatory effects. The analyzed iridoids and RA, individually or in mixtures, have the potential to be used in the pharmaceutical industry as potent antimicrobials, and in the food industry to increase the shelf life and safety of food products.

Microalgae show significant biotechnological potential for remediation of wastewaters polluted with metals, including nickel. Ni enters aquatic ecosystems from fuel combustion, smelting, mining, and electroplating endeavors, and represents a particularly important problem for the access to safe drinking water. It is noteworthy that microalgae utilize Ni as a co-factor of urease. Herein we analyzed the impact of high levels of Ni(II) on freshwater microalga Chlorella sorokiniana in the stationary phase of culture growth. In the concentration range 0.5 to 30 mM, Ni induced a drop in cell density and biomass after 7 days of incubation, whereas significant negative impact was present as early as 2 days for 30 mM. High concentrations also induced a decrease in the level of chlorophylls (a and b) in biomass. In contrast, microalgae that were exposed to moderate stress (1 and 2 mM of Ni) appear to increase photosynthetic activity, indicating the activation of some adaptive mechanism. Scanning electron microscopy showed that C. sorokiniana does not release significant amounts of mucilage polymers in response to Ni, in contrast to the response to some other metals 3. Further, Ni at 1 mM induced an increase in the intracellular production of reactive oxygen species, which appears to reach a plateau after 1 h. The concentration of free thiol groups showed a gradual drop during 24 h of incubation with the same Ni concentration. On the other hand, the decrease in the concentration of reduced glutathione that was observed after 1 h was reversible for 1 mM Ni at 24 h. Ni at 5 mM had a more lasting impact on glutathione. It is important to stress out that the concentrations of reduced glutathione and total glutathione showed very similar trends. This implies that free thiols and reduced glutathione may be ‘lost’ due to glutathionylation of proteins and synthesis of phytochelatins. Our results demonstrate that the response of C. sorokiniana to high Ni levels involves multiple compon

Microalgae can be implemented in the remediation of mining and other metal-rich wastewaters as pioneer species. On the other hand, metals affect aquatic ecosystems through the negative impact on microalgae which are the primary producers of O2 and biomass. Redox processes represent an important component of the mechanisms of interaction of microalgae with transition metals. We analyzed the redox changes in Chlorella sorokiniana culture that are induced by high levels of Mn(II). Mn is the key metal pollutant, with five main oxidation forms that can bind to a variety of different ligands. Mn (1 mM) induced a significant increase in the intracellular production of reactive oxygen species. The boost appears to show two phases – the first is very fast (observed after 15 min), whereas the second starts after 1 h reaching a plateau at 24 h. The concentration of reduced thiols, which represent important targets of oxidation, appears to parallel this trend. Total glutathione concentration shows a drop at 1 h and recovery at 24 h. This implicates that either a glutathionylation of proteins or a synthesis of phytochelatins - sulfur-rich short-chain peptides that sequester metals, takes place early in the response to Mn. Further, FTIR analysis showed that Mn induced a decrease of C=C levels and CH2/CH3 ratio implicating increased lipid peroxidation. Finally, Mn ions that were accumulated in the cells were extracted with nitric oxide and analyzed by cyclic voltammetry. Two redox forms were detected - Mn(II) and Mn(IV). The latter appears to prevail at higher manganese concentrations and longer periods of incubation. These results demonstrate that redox response of C. sorokiniana to high Mn levels involves at least two phases. Initially, Mn(II) enters the cells and induces pro-oxidative changes that are mitigated by glutathione-based antioxidative defense. Later on, redox homeostasis is reestablished with concomitant inactivation of Mn in the more stable redox form.

Biliverdin (BV), proizvod katabolizma hem grupe, u interakciji sa prelaznim metalima može da gradi komplekse, ali detalji ovih interakcija u fiziološkim uslovima nisu dovoljno opisani. Ovde su ispitane koordicione/redoks interakcije BV sa jonima Cu(II) u fosfatnom puferu na pH 7,4, koristeći ultraljubičastu-vidljivu spektrofotometriju, masenu spektrometriju, ramansku spektroskopija, metodu nuklearne magnetne rezonacije (NMR), metodu elektronske paramagnetne rezonacije (EPR), fluorimetrijske i elektrohemijske metode. Pokazano je da BV formira stabilan koordinacioni kompleks sa jonima bakra u stehiometriji 1:1. Struktura BV je više planarna i energijski stabilna u kompleksu. Kompleks je pokazao jake paramagnetne efekte koji su pripisani nesparenom delokalizovanom elektronu. Delokalizovani elektron može poticati iz BV ili od jona Cu(II), pa je kompleks sastavljen ili od katjona BV radikala i jona Cu(I) ili od anjona BV radikala i jona Cu(III). Kompleks je podložan oksidaciji samo u prisustvu O2 i višku jona Cu(II), ili jakog oksidacionog sredstva, dok je otporan na redukciona sredstva. Biološki efekti stabilnog metalokompleksa BV koji sadrže delokalizovani nesparni elektron zahtevaju dalje istraživanje. Ovo može dati odgovor na dugogodišnje pitanje razloga ulaganje velike količine energije u katabolizam BV, koji sam po sebi predstavlja relativno bezopasan molekul..., Biliverdin (BV) is one of the products of heme catabolism that is known to interact with redox-active metals. (Patho)physiological roles of BV and details of its interactions with metals under physiological conditions, which may be interconnected, are not fully understood. In the current work, we examined coordinate and redox interactions of BV with copper ions (Cu(II)). The interactions were exaimined under conditions that mimic physiological settings - in the phosphate buffer with pH 7.4. A battery of physico-chemical methods were applied, including UV-VIS spectrophotometry, fluorescence spectroscopy, nuclear magnetic resonance spectroscopy (NMR), mass spectrometry, Raman spectroscopy, electron paramagnetic resonance (EPR), and electrochemical methods. BV and copper formed a stable coordination complex with 1 : 1 stoichiometry, with BV showing a more planar structure and energetic stability in the complex. The complex showed strong paramagnetic effects, most likely due to a delocalized unpaired electron. The delocalized e- may come from copper ion or BV. The complex is formally composed of BV radical cation and Cu(I) ion or of BV radical anion and Cu(III) ion. The complex underwent oxidation only in the presence of both O2 and an excess of Cu(II) ion, or a strong oxidizing agent, and it was resistant to reducing agents. The biological effects of the stable BV metallocomplex containing a delocalized unpaired electron should be further examined, and may provide an answer to the long-standing question of high energy investment in the catabolism of BV, which represents a relatively harmless molecule per se...

Upon release in response to stress, epinephrine (Epi) may interact with labile iron pool in human plasma with potentially important (patho)physiological consequences. We have shown that Epi and Fe3+ build stable 1:1 high-spin bidentate complex at physiological pH, and that Epi does not undergo degradation in the presence of iron. However, the interactions of Epi with the more soluble Fe2+, and the impact of iron on biological activity of Epi are still not known. Herein we showed that Epi and Fe2+ build colorless complex which is stable under anaerobic conditions. In the presence of O2, Epi promoted the oxidation of Fe2+ and the formation of Epi-Fe3+ complex. Cyclic voltammetry showed that mid-point potential of Epi-Fe2+ complex is very low (−582 mV vs. standard hydrogen electrode), which explains catalyzed oxidation of Fe2+. Next, we examined the impact of iron binding on biological performance of Epi using patch clamping in cell culture with constitutive expression of adrenergic receptors. Epi alone evoked an increase of outward currents, whereas Epi in the complex with Fe3+ did not. This implies that the binding of Epi to adrenergic receptors and their activation is prevented by the formation of complex with iron. Pro-oxidative activity of Epi-Fe2+ complex may represent a link between chronic stress and cardiovascular problems. On the other hand, labile iron could serve as a modulator of biological activity of ligands. Such interactions may be important in human pathologies that are related to iron overload or deficiency.

Microalgae have evolved mechanisms to respond to changes in copper ion availability, which are very important for normal cellular function, to tolerate metal pollution of aquatic ecosystems, and for modulation of copper bioavailability and toxicity to other organisms. Knowledge and application of these mechanisms will benefit the use of microalgae in wastewater processing and biomass production, and the use of copper compounds in the suppression of harmful algal blooms. Here, using electron microscopy, synchrotron radiation-based Fourier transform infrared spectroscopy, electron paramagnetic resonance spectroscopy, and X-ray absorption fine structure spectroscopy, we show that the microalga Chlorella sorokiniana responds promptly to Cu2+ at high non-toxic concentration, by mucilage release, alterations in the architecture of the outer cell wall layer and lipid structures, and polyphosphate accumulation within mucilage matrix. The main route of copper detoxification is by Cu2+ coordination to polyphosphates in penta-coordinated geometry. The sequestrated Cu2+ was accessible and could be released by extracellular chelating agents. Finally, the reduction in Cu2+ to Cu1+ appears also to take place. These findings reveal the biochemical basis of the capacity of microalgae to adapt to high external copper concentrations and to serve as both, sinks and pools of environmental copper.

Biliverdin (BV), a product of heme catabolism, is known to interact with transition metals, but the details of such interactions under physiological conditions are scarce. Herein, we examined coordinate/redox interactions of BV with Cu2+ in phosphate buffer at pH 7.4, using spectrophotometry, HESI-MS, Raman spectroscopy, 1 H NMR, EPR, fluorimetry, and electrochemical methods. BV formed a stable coordination complex with copper in 1 : 1 stoichiometry. The structure of BV was more planar and energetically stable in the complex. The complex showed strong paramagnetic effects that were attributed to an unpaired delocalized e−. The delocalized electron may come from BV or Cu2+, so the complex is formally composed either of BV radical cation and Cu1+ or of BV radical anion and Cu3+. The complex underwent oxidation only in the presence of both O2 and an excess of Cu2+, or a strong oxidizing agent, and it was resistant to reducing agents. The biological effects of the stable BV metallocomplex containing a delocalized unpaired electron should be further examined, and may provide an answer to the long-standing question of high energy investment in the catabolism of BV, which represents a relatively harmless molecule per se.

Biliverdin (BV), a product of heme catabolism, is known to interact with transition metals, but the details of such interactions under physiological conditions are scarce. Herein, we examined coordinate/redox interactions of BV with Cu 2+ in phosphate buffer at pH 7.4, using spectrophotometry, HESI-MS, Raman spectroscopy, 1 H NMR, EPR, fluorimetry, and electrochemical methods. BV formed a stable coordination complex with copper in 1:1 stoichiometry. The structure of BV was more planar and energetically stable in the complex. The complex showed strong paramagnetic effects that were attributed to an unpaired delocalized e - . The delocalized electron may come from BV or Cu 2+ , so the complex is formally composed either of BV radical cation and Cu 1+ or of BV radical anion and Cu 3+ . The complex underwent oxidation only in the presence of both O 2 and an excess of Cu 2+ , or a strong oxidizing agent, and it was resistant to reducing agents. The biological effects of the stable BV metallocomplex containing a delocalized unpaired electron should be further examined, and may provide an answer to the long-standing question of high energy investment in the catabolism of BV, which represents a relatively harmless molecule per se., Accepted Author Manuscript, BT/Biocatalysis

Biliverdin (BV) structure was analyzed by using NMR techniques and unrestricted density function theory simulations to explain the incapacity of BV to build coordination complex(es) with Cu2+ in dimethyl sulfoxide, which was confirmed by UV-Vis, EPR and NMR spectroscopy. NMR showed that N atoms are protonated in all four pyrrole rings. The structure is stabilized by two hydrogen bonds between NH moieties and carbonyl oxygens from opposite terminal pyrrole rings, and by the bending of propionyl chain with carboxyl group out of the plain toward central position of BV. The simulations of deprotonated BV, which builds copper complexes in water and chloroform as described previously, showed a different conformation and organization of hydrogen bonds. Taking into account that deprotonation represents a critical step in coordinate bonds formation, the protonation of an additional N atom may represent a key difference between the interactions of BV with copper in different solvents.

Biliverdin (BV), a product of heme catabolism, is known to interact with transition metals, but the details of such interactions under physiological conditions are scarce. Herein, we examined coordinate/redox interactions of BV with Cu2+ in phosphate buffer at pH 7.4, using spectrophotometry, HESI-MS, Raman spectroscopy, H-1 NMR, EPR, fluorimetry, and electrochemical methods. BV formed a stable coordination complex with copper in 1:1 stoichiometry. The structure of BV was more planar and energetically stable in the complex. The complex showed strong paramagnetic effects that were attributed to an unpaired delocalized e(-). The delocalized electron may come from BV or Cu2+, so the complex is formally composed either of BV radical cation and Cu1+ or of BV radical anion and Cu3+. The complex underwent oxidation only in the presence of both O-2 and an excess of Cu2+, or a strong oxidizing agent, and it was resistant to reducing agents. The biological effects of the stable BV metallocomplex containing a delocalized unpaired electron should be further examined, and may provide an answer to the long-standing question of high energy investment in the catabolism of BV, which represents a relatively harmless molecule per se.

The structure of biliverdin (BV) and its complexion capacity with Cu2+ in dependence of solvent choice has been examined. We first examined coordinate/redox interactions of BV with Cu2+ in phosphate buffer at pH 7.4, using spectrophotometry, HESI-MS, Raman spectroscopy, 1HNMR, EPR, fluorimetry, and electrochemical methods. BV formed a stable coordination complex with copper in 1 : 1 stoichiometry. The structure of BV was more planar and energetically stable in the complex. The complex showed strong paramagnetic effects that were attributed to an unpaired delocalized e−. The delocalized electron may come from BV or Cu2+, so the complex is formally composed either of BV radical cation and Cu1+ or of BV radical anion and Cu3+. The complex underwent oxidation only in the presence of both O2 and an excess of Cu2+, or a strong oxidizing agent, and it was resistant to reducing agents. The biological effects of the stable BV metallocomplex containing a delocalized unpaired electron should be further examined, and may provide an answer to the longstanding question of high energy investment in the catabolism of BV, which represents a relatively harmless molecule per se. The complex possibility was quite different after changing the solvent. BV structure in DMSO was analyzed by using NMR techniques and unrestricted density function theory simulations to explain the incapacity of BV to build coordination complex(es) with Cu2+ in dimethyl sulfoxide, which was confirmed by UV-Vis, EPR and NMR spectroscopy. NMR showed that N atoms of BV are protonated in all four pyrrole rings. The structure is stabilized by two hydrogen bonds between NH moieties and carbonyl oxygens from opposite terminal pyrrole rings, and by the bending of propionyl chain with carboxyl group out of the plain toward central position of BV. The simulations of deprotonated BV, which builds copper complexes in water and chloroform as described previously, showed a different conformation and organization of hydrogen

Radiation hormesis is a biological phenomenon whereby a relatively low doses of radiation induce beneficial effects on biological parameters, such as viability, growth, and metabolic profile 1. A number of studies have documented stimulatory effects of low-dose X- and gamma radiation on plant growth 2. However, radiation hormesis has not been investigated in microalgae which are becoming a highly relevant source of lipid-derived biofuels. The aim of this study was to establish the effects of low-dose X-irradiation on biomass and lipid production in Chlorella sorokiniana. The microalgae cultures were exposed to total doses of 10, 20 or 50 Gy at different irradiation rates (0.565; 2.290 and 5.057 Gy/min). Absorbed doses corresponded to ~10% of the total dose, as determined by dosimetry. Lipid content was determined 30 days after irradiation using Nile Red fluorescence assay with excitation/emission maxima at 530/570 nm (total lipids), 480/575 nm (neutral lipids) and 549/628 nm (polar lipids) 3,4. Lipid content was normalized to dry biomass.

Introduction. We tested whether the microclimate under grey hail protection nets (HPN) had an influence on the phenology, vegetative and reproductive potential, and fruit quality traits of 'Duke' highbush blueberry (Vaccinium corymbosum L.) in two consecutive years (2013, 2014). Materials and methods. Light intensity, air temperature and relative humidity were evaluated together with biological properties (flowering and ripening time, bush height and volume, number of flowers and fruits per bush, fruit set percentage, and yield per bush), and fruit quality including biometrical (fruit weight, index of fruit shape and number of seeds per fruit) and nutritional (soluble solid content SSC, titratable acidity - TA, total anthocyanins content - TACY, total phenolic content - TPC and total antioxidant capacity - TAC) parameters. Results and discussion. In both seasons, light was reduced by 5-20% under HPN, whereas daily maximum temperature was 2.4 degrees C higher in the open field (OF). This caused two days earlier ripening time in OF compared to the HPN. The increased yield per bush under HPN was the result of larger fruit weight. SSC were unaffected by the net in 2013 while in 2014 significantly higher value of SSC was registered under HPN compared to OF. Although apparent differences in TACY and TPC were not observed between the treatments, TAC significantly increased under HPN in 2014. Conclusion. The use of grey HPN could be considered as a suitable alternative for the hail protection of highbush blueberry ensuring an increased fruit production without negative effects on their quality.

Introduction. We tested whether the microclimate under grey hail protection nets (HPN) had an influence on the phenology, vegetative and reproductive potential, and fruit quality traits of 'Duke' highbush blueberry (Vaccinium corymbosum L.) in two consecutive years (2013, 2014). Materials and methods. Light intensity, air temperature and relative humidity were evaluated together with biological properties (flowering and ripening time, bush height and volume, number of flowers and fruits per bush, fruit set percentage, and yield per bush), and fruit quality including biometrical (fruit weight, index of fruit shape and number of seeds per fruit) and nutritional (soluble solid content SSC, titratable acidity - TA, total anthocyanins content - TACY, total phenolic content - TPC and total antioxidant capacity - TAC) parameters. Results and discussion. In both seasons, light was reduced by 5-20% under HPN, whereas daily maximum temperature was 2.4 degrees C higher in the open field (OF). This caused two days earlier ripening time in OF compared to the HPN. The increased yield per bush under HPN was the result of larger fruit weight. SSC were unaffected by the net in 2013 while in 2014 significantly higher value of SSC was registered under HPN compared to OF. Although apparent differences in TACY and TPC were not observed between the treatments, TAC significantly increased under HPN in 2014. Conclusion. The use of grey HPN could be considered as a suitable alternative for the hail protection of highbush blueberry ensuring an increased fruit production without negative effects on their quality.