Calcium-enriched biochar modulates cadmium uptake depending on external cadmium dose.

The impact of calcium-enriched biochar (BC, containing Ca, Al, Fe and P as dominant elements in the range of 6.9-1.3% with alkaline pH) obtained from sewage sludge (0.1 or 0.5% in the final soil) on cadmium-induced toxicity (final dose of 1.5 mg Cd/kg in control and 4.5 or 16.5 mg Cd/kg soil in low and high Cd treatment) was tested in medicinal plant Matricaria chamomilla. Low Cd dose had typically less negative impact than high Cd dose at the level of minerals and metabolites and the effect of BC doses often differed. Contrary to expectations, 0.5% BC with a high Cd dose increased Cd accumulation in plants about 2-fold. This was reflected in higher signals of reactive oxygen species, but especially the high dose of BC increased the amount of antioxidants (ascorbic acid and non-protein thiols), minerals and amino acids in shoots and/or roots and usually mitigated the negative effect of Cd. Surprisingly, the relationship between BC and soluble phenols was negative at high BC + high Cd dose, whereas the effect of Cd and BC on organic acids (mainly tartaric acid) differed in shoots and roots. Interestingly, BC alone applied to the control soil (1.5 mg total Cd/kg) reduced the amount of Cd in the plants by about 30%. PCA analyses confirmed that metabolic changes clearly distinguished the high Cd + high BC treatment from the corresponding Cd/BC treatments in both shoots and roots. Thus, it is clear that the effect of biochar depends not only on its dose but also on the amount of Cd in the soil, suggesting the use of Ca-rich biochar both for phytoremediation and safer food production.

Fatty Acid Supplementation Affects Skin Wound Healing in a Rat Model.

Polyunsaturated fatty acids (PUFA) play an important role in reparative processes. The ratio of PUFAs n-3 to n-6 may affect wound healing. The study aimed to evaluate the effect of dietary supplementation with n-3 and n-6 PUFA in two proportions on skin wounds in laboratory rats. Adult male Wistar rats received 20% fat emulsion with a ratio of 1.4:1 (group A) or 4.3:1 (group B) for n-3:n-6 PUFAs at a daily dose of 1 mL/kg. The control group received water under the same conditions. The animals were supplemented a week before and a week after the skin excision performed on the back. The level of wound closure, various parameters of oxidative stress, and plasma fatty acids composition were evaluated. Wound tissue samples were examined by electron microscopy. The administration of fat emulsions led to significant changes in plasma polyunsaturated fatty acid composition. The increased production of reactive nitrogen species, as well as more numerous newly formed blood vessels and a greater amount of highly organized collagen fibrils in both groups A and B may indicate more intensive healing of the skin wound in rats supplemented with polyunsaturated fatty acids in high n-3:n-6 ratio.

Nitrogen modulates strontium uptake and toxicity in Hypericum perforatum plants.

Strontium is an unavoidable element occurring in plants due to its abundance in the soil and similarity with calcium. To mimic natural conditions, impacts of additional inorganic (nitrate) or organic (urea and allantoin) nitrogen sources (1 mM of each N form in addition to 3.53 mM N in the basic cultivation solution) or N deficit on strontium-induced changes (100 µM Sr) in the widely used medicinal plant Hypericum perforatum L. were studied. Though various effects of Sr on primary (stimulation of amino acids but depression of most Krebs acids, ascorbic acid and thiols) and secondary metabolites (stimulation of phenols but no change of pseudo/hypericin) or mineral elements were observed (reduction of Ca amount in both shoots and roots), organic N forms often mitigated negative action of Sr or even combined stimulatory impact was observed. Organic N forms also elevated shoot accumulation of Sr while N deficit reduced it. Additional N forms, rather than Sr itself, modulated reactive oxygen species and nitric oxide formation in the root tissue. Germination experiment showed no toxicity of Sr to H. perforatum up to 1 mM Sr and even stimulated accumulation of amino acids and phenols, indicating similar ontogenetic-related responses.

Calcium signaling affects migration and proliferation differently in individual cancer cells due to nifedipine treatment.

Several papers have reported that calcium channel blocking drugs were associated with increased breast cancer risk and worsened prognosis. One of the most common signs of breast tumors is the presence of small deposits of calcium, known as microcalcifications. Therefore, we studied the effect of dihydropyridine nifedipine on selected calcium transport systems in MDA-MB-231 cells, originating from triple negative breast tumor and JIMT1 cells that represent a model of HER2-positive breast cancer, which possesses amplification of HER2 receptor, but cells do not response to HER2 inhibition treatment with trastuzumab. Also, we compared the effect of nifedipine on colorectal DLD1 and ovarian A2780 cancer cells. Both, inositol 1,4,5-trisphosphate receptor type 1 (IP3R1) and type 1 sodium calcium exchanger (NCX1) were upregulated due to nifedipine in DLD1 and A2780 cells, but not in breast cancer MDA-MB-231 and JIMT1 cells. On contrary to MDA-MB-231 and JIMT1 cells, in DLD1 and A2780 cells nifedipine induced apoptosis in a concentration-dependent manner. After NCX1 silencing and subsequent treatment with nifedipine, proliferation was decreased in MDA-MB-231, increased in DLD1 cells, and not changed in JIMT1 cells. Silencing of IP3R1 revealed increase in proliferation in DLD1 and JIMT1 cells, but caused decrease in proliferation in MDA-MB-231 cell line after nifedipine treatment. Interestingly, after nifedipine treatment migration was not significantly affected in any of tested cell lines after NCX1 silencing. Due to IP3R1 silencing, significant decrease in migration occurred in MDA-MB-231 cells after nifedipine treatment, but not in other tested cells. These results support different function of the NCX1 and IP3R1 in the invasiveness of various cancer cells due to nifedipine treatment.

Nitrogen nutrition modulates oxidative stress and metabolite production in Hypericum perforatum.

Impact of various nitrate concentrations (14.12 mM, 3.53 mM, no nitrate) or ammonium presence (14.12 mM) on physiological and metabolic changes in Hypericum perforatum after 14 days of cultivation was monitored. Nitrate deficiency suppressed growth of shoots but stimulated root growth while ammonium suppressed root growth: concomitant changes of ascorbic acid and glutathione supported these growth changes, e.g., unaltered level in roots under nitrate deficiency but depleted in ammonium treatment. Soluble proteins and water content were more suppressed by nitrate deficiency but total ROS, nitric oxide formation, and antioxidative enzyme activities (APX and SOD) indicate higher sensitivity of plants to ammonium. Though both extreme treatments (NO3- deficiency or ammonium) stimulated accumulation of total soluble phenols and affected PAL activity (in comparison with full or 1/4× nitrate dose), major phenols (chlorogenic acid and three flavonoids) were elevated mainly by NO3- deficiency. At the level of specific metabolites, NO3- deficiency had stimulatory impact on pseudohypericin (but not hypericin) content while hyperforin decreased. Expression of earlier putative gene of hypericin biosynthesis (hyp-1) showed rather partial correlation with pseudohypericin amount. Data indicate that depletion of NO3- is useful to obtain Hypericum plants with higher amount of health-positive secondary metabolites.

Metabolic responses of Ulva compressa to single and combined heavy metals.

Accumulation of metals and metabolic responses were studied for two Cd and Cu concentrations (1 and 10 µM) either alone or as a combination in marine macroalga after 7 days of exposure. Cd accumulated more at a low dose (115 µg of Cd/g DW) but Cu at a high dose (378 µg of Cu/g DW); Cu suppressed Cd accumulation (by 57%). Na and Zn levels were unaffected, but higher metal doses depleted K and Ca levels. Higher metal concentrations strongly stimulated reactive oxygen species and depleted nitric oxide (NO) formation, but differences between the action of Cd and Cu were not extensive. Higher metal doses increased cell wall thickness with a potential relation to NO signal that is visible mainly in the apoplast in those treatments. A higher Cu dose depleted proline, ascorbic acid, and phenol levels more than Cd, whereas Cd elevated nonprotein thiols and ascorbic acid in combined treatments. An eventual role of malic or citric acid in metal chelation was not evident: malic acid level decreased in all treatments. The total content of fatty acids reached 16.7 mg/g DW in control with the quantitative order of PUFAs > SFAs > MUFAs; palmitic, vaccenic, linoleic, and α-linolenic acids were the major compounds. Cu was more toxic for fatty acids than Cd (even at 1 µM); mainly, PUFA levels strongly decreased (from 43% of total acids in control to 28.9% and 5.4% at 1 and 10 µM Cu treatment, respectively). Results are precisely and critically discussed in relation to limited literature focused on macroalgae, and a comparison with microalgae is also provided.

Lanthanum rather than cadmium induces oxidative stress and metabolite changes in Hypericum perforatum.

Physiology, oxidative stress and production of metabolites in Hypericum perforatum exposed to moderate Cd and/or La concentration (10 µM) were studied. La evoked increase in reactive oxygen species, malondialdehyde and proline but suppressed growth, tissue water content, glutathione, ascorbic acid and affected mineral nutrient contents more than Cd while the impact of Cd+La was not synergistic. Similar trend was observed at the level of superoxide dismutase gene expression. Shoot Cd amount increased in Cd+La while only root La increased in the same treatment. Extensive quantification of secondary metabolites revealed that La affected phenolic acids more pronouncedly than Cd in shoots and roots. Flavonols were suppressed by La that could contribute to the appearance of oxidative damage. Procyanidins increased in response to La in the shoots but decreased in the roots. Metabolic responses in Cd+La treatment resembled those of La treatment (almost identically in the roots). Phenylalanine ammonia-lyase activity was mainly suppressed by La. The presence of La also depleted amount of hypericin and expression of its putative gene (hyp-1) showed similar trend but accumulation of hyperforin increased under Cd or La excess. Clear differences in the stem and root anatomy in response to Cd or La were also found. Overall, H. perforatum is La-sensitive species and rather Cd ameliorated negative impact of La.

Turkish Scorzonera Species Extracts Attenuate Cytokine Secretion via Inhibition of NF-κB Activation, Showing Anti-Inflammatory Effect in Vitro.

Scorzonera species are used in different folk medicines to combat many diseases, including the illnesses connected with inflammation. Previous experiments showed anti-inflammatory activity of Scorzonera extracts in vivo. S. latifolia, S. cana var. jacquiniana, S. tomentosa, S. mollis ssp. szowitsii, S. eriophora, S. incisa, S. cinerea, and S. parviflora extracts were, therefore, evaluated for their inhibitory activities of TNF-α and IL-1ß production, and NF-κB nuclear translocation in THP-1 macrophages. The HPLC analysis was carried out to elucidate and to compare the composition of these extracts. Major compounds of the tested extracts have been isolated using different chromatographic techniques and further tested for their inhibitory activities on TNF-α and IL-1ß production. Several extracts showed promising anti-inflammatory activity in these in vitro tests. Results of HPLC analysis revealed chlorogenic acid as a compound present in all tested extracts. Hyperoside, quercetin-3-O-ß-d-glucoside and rutin were also present in varying amount in some Scorzonera species analyzed. Furthermore, eight phenolics which were identified as quercetin-3-O-ß-d-glucoside (1), hyperoside (2), hydrangenol-8-O-glucoside (3), swertisin (4), 7-methylisoorientin (5), 4,5-O-dicaffeoyl-quinic acid (6), 3,5-di-O-caffeoyl-quinic acid (7), and chlorogenic acid (8) have been isolated as major phenolic compounds of the tested extracts and, together with eight terpenoids (9-16) previously obtained from different Scorzonera species, have been tested for the inhibition of TNF-α production, unfortunately with no activity comparable with standard.

PAL inhibitor evokes different responses in two Hypericum species.

Accumulation of secondary metabolites (general phenols, naphthodianthrones and phloroglucinol hyperforin) in Hypericum perforatum and Hypericum canariense after application of the inhibitor (2-aminoindane-2-phosphonic acid, AIP) of the pivotal enzyme of general phenylpropanoid pathway (phenylalanine ammonia-lyase, PAL) was studied. Shoots of H. perforatum revealed more expressive growth depression, concomitantly with the inhibition of PAL activity (-60%) and decrease in soluble phenols and individual phenolic acids in response to AIP. Hypericins (hypericin, pseudohypericin and protohypericin) decreased while hyperforin increased in AIP-cultured H. perforatum. On the contrary, growth changes, decreases in soluble phenols and individual phenolic acids were less-visible in H. canariense. This was also reflected in restoration of PAL activity (+330%) and selected flavonoids even increased. Hypericins and hyperforin were present in several orders of magnitude lower amounts in comparison with H. perforatum. Increase in proline indicates potential compensatory antioxidative mechanism if phenols are depleted. Microscopy revealed also differences in secondary xylem formation and lignification between species after exposure to AIP.

Mathematical evaluation of the amino acid and polyphenol content and antioxidant activities of fruits from different apricot cultivars.

Functional foods are of interest because of their significant effects on human health, which can be connected with the presence of some biologically important compounds. In this study, we carried out complex analysis of 239 apricot cultivars (Prunus armeniaca L.) cultivated in Lednice (climatic area T4), South Moravia, Czech Republic. Almost all previously published studies have focused only on analysis of certain parameters. However, we focused on detection both primary and secondary metabolites in a selection of apricot cultivars with respect to their biological activity. The contents of thirteen biogenic alpha-L-amino acids (arginine, asparagine, isoleucine, lysine, serine, threonine, valine, leucine, phenylalanine, tryptophan, tyrosine, proline and alanine) were determined using ion exchange chromatography with UV-Vis spectrometry detection. Profile of polyphenols, measured as content of ten polyphenols with significant antioxidant properties (gallic acid, procatechinic acid, p-aminobenzoic acid, chlorogenic acid, caffeic acid, vanillin, p-coumaric acid, rutin, ferrulic acid and quercetrin), was determined by high performance liquid chromatography with spectrometric/electrochemical detection. Moreover, content of total phenolics was determined spectrophotometrically using the Folin-Ciocalteu method. Antioxidant activity was determined using five independent spectrophotometric methods: DPPH assay, DMPD method, ABTS method, FRAP and Free Radicals methods. Considering the complexity of the obtained data, they were processed and correlated using bioinformatics techniques (cluster analysis, principal component analysis). The studied apricot cultivars were clustered according to their common biochemical properties, which has not been done before. The observed similarities and differences were discussed.

Effects of various doses of selenite on stinging nettle (Urtica dioica L.).

The aim of this study was to investigate the effects of selenium (Se) on the growth, accumulation and possible mechanisms of Se transport in certain parts (roots, leaves, stamp and apex) of nettle (Urtica dioica L.) plants. Se was supplemented by one-shot and two repeated doses to the soil (2.0 and 4.0 mg Se per kg of substrate). Selenium content in roots increased linearly with dose and was significantly higher compared to other plant parts of interest. However, growth of the above-ground parts of plant as well as roots was slightly inhibited with increasing selenium concentration in comparison to the untreated plants. The content of phytochelatin2, a low molecular mass peptide containing a sulfhydryl group, correlated well with the Se content. This suggests a possible stimulation of synthesis of this plant peptide by Se.

Evaluation of cytotoxic activity of Schisandra chinensis lignans.

Using exhaustive chromatographic separation we have isolated (-)-tigloyl-deangeloyl-gomisin F as a novel dibenzocyclooctadiene lignan from schisandra chinensis. With the help of HPLC, we further isolated (+)-schisandrin, (+)-deoxyschisandrin, (+)-γ-schisandrin, (-)-gomisin J, (+)-gomisin A, (-)-gomisin N, (-)-tigloyl-gomisin P, (-)-wuweizisu C, (-)-gomisin D, rubrisandrin A, (-)-gomisin G, (+)-gomisin K (3) and (-)-schisantherin C. A full NMR description of (-)-schisantherin C was carried out with the aim to confirm previous reports of its structure. Compounds isolated were identified on the basis of UV, IR, (1)H- and (13)C-NMR and MS. The cytotoxicity of lignans was tested for the BY-2 cell line alone and as a synergistic effect with the cytotoxic agent camptothecin. Lignans showed various toxicity and synergistic and antagonistic effects on camptothecin-induced cytotoxicity. Cytotoxicity against colon cancer cell line LoVo was also tested.

Cytotoxic activity of C-geranyl compounds from Paulownia tomentosa fruits.

The newly discovered 5,7-dihydroxy-6-geranylchromone ( 1) was isolated from PAULOWNIA TOMENTOSA fruit and subsequently characterized. The structure of the isolated compound was elucidated on the basis of extensive NMR experiments including HMQC, HMBC, COSY, and NOESY, as well as HR-MS, IR, and UV. The cytotoxicity of 1 was evaluated using a plant cell model represented by tobacco BY-2 cells. The other phytoconstituents ( 2 - 8) previously isolated from P. TOMENTOSA were similarly evaluated together with the known flavanones 10 and 11. The cytotoxicity (human erythro-leukaemia cell line K562) and activity on erythroid differentiation of compounds 2 - 9 and 12 and 13 have also been evaluated. Acteoside ( 2) was determined to be the most toxic of the compounds tested on BY-2 cells, diplacone ( 6) on the K562 cell line. Some aspects of the relationship between the flavanone skeleton substitution and the metabolic activation necessary for a toxic effect are discussed.

Electrochemical Determination of Low Molecular Mass Thiols Content in Potatoes (Solanum tuberosum) Cultivated in the Presence of Various Sulphur Forms and Infected by Late Blight (Phytophora infestans).

In the present paper potato plants were cultivated in the presence of ammonium sulphate or elemental sulphur supplementation into the soil to reveal the effects of different sulphur forms on content of nitrogen, phosphorus, potassium, calcium, magnesium and sulphur, and yield of tubers. During the investigation of the influence of different sulphur forms on yield of potato tubers we did not observe significant changes. Average weight of tubers of control plants per one experimental pot was 355 g. Application of sulphur in both forms resulted in moderate potato tubers weight reduction per one experimental pot compared to control group; average value ranged from 320 to 350 g per one experimental pot. Further we treated the plants with two different supplementation of sulphur with cadmium(II) ions (4 mg of cadmium(II) acetate per kilogram of the soil). The significantly lowest cadmium content (p < 0.05) was determined in tissues of plants treated with the highest dosage of elemental sulphur (0.64 mg Cd/kg) compared to control plants (0.82 mg Cd/kg). We also aimed our attention on the cadmium content in proteins, lipids or soluble carbohydrates and ash. Application of sulphate as well as elemental sulphur resulted in significant cadmium content reduction in lipid fraction compared to control plants. In addition to this we quantified content of low molecular mass thiols in potatoes tissues. To determine the thiols content we employed differential pulse voltammetry Brdicka reaction. After twelve days of the treatment enhancing of thiols level was observed in all experimental groups regardless to applied sulphur form and its concentration. Finally we evaluated the effect of sulphur supplementation on Phytophora infestans infection of potato plants.