Metabolism of Mycosporine-Glutamicol in the Lichen Cladonia arbuscula subsp. squarrosa under Seasonal Changes and Elevated Exposure to UV-B or PAR Irradiation.

Cladonia arbuscula in its environmental niches is regularly affected by daily and annual variations in solar radiation. Mycosporine-glutamicol, Myc-Glu(OH), which it synthesizes, may act as a significant cellular UV-protector. Therefore, we studied this compound concentration in lichen thalli concerning seasonal changes and increased exposure to UV-B and photosynthetically active radiation (PAR) with/without simultaneous CO2 deprivation. Myc-Glu(OH) occurred year-round and exhibited a strong seasonality. The most crucial role in the control of its synthesis played UV-B radiation, although its high concentration was also found after PAR irradiation at 1000 µmol m-2 s-1. As PAR intensity increased to 2000 µmol m-2 s-1, the rate of Myc-Glu(OH) synthesis slowed down. In turn, under dark/PAR irradiation with simultaneous deprivation of CO2 in the atmosphere surrounding C. arbuscula and during darkness with continuous access to atmospheric CO2, its production was insignificant. Obtained data confirmed that Myc-Glu(OH) plays an important role in protecting C. arbuscula from UV damage and favours its adaptation to environmental stress in its natural habitat. They also suggest that its synthesis is a synergism of multiple factors. Consequently, further studies should focus on their evaluation and the identification of a lichen partner actively involved in Myc-Glu(OH) biogenesis.

Inhibition of growth rate and cylindrospermopsin synthesis by Raphidiopsis raciborskii upon exposure to macrophyte Lemna trisulca (L).

Impact of macrophyte Lemna trisulca on the growth rate and synthesis of cylindrospermopsin (CYN) by cyanobacterium Raphidiopsis raciborskii was determined. The presence of L. trisulca inhibited the biomass accumulation of the cyanobacterium by 25% compared to the control during co-cultivation. The simultaneous cultivation of these organisms slightly affected the inhibition of macrophyte growth rate by 5.5% compared to the control. However, no morphological changes of L. trisulca after incubation with cyanobacteria were observed. It was also shown that the long-term (35 days) co-cultivation of R. raciborskii and L. trisulca led to a decrease in CYN concentration in media and cyanobacterial cells by 32 and 38%, respectively, compared to the values obtained for independent cultivation of cyanobacterium. Excessive absorption of phosphate ions by L. trisulca from the medium compared to nitrate ions led to a significant increase in the nitrate:phosphate ratio in the media, which inhibits the development of cyanobacterium. The obtained results indicate that L. trisulca in the natural environment may affect the physiology of cyanobacteria. The presented study is the first assessment of the allelopathic interaction of macrophyte and R. raciborskii.

Effects of cylindrospermopsin, its decomposition products, and anatoxin-a on human keratinocytes.

Toxins produced by cyanobacteria (cyanotoxins) are among the most dangerous natural compounds. In recent years, there have been many published papers related to the toxic alkaloids cylindrospermopsin (CYN) and anatoxin-a (ANTX-a), which are synthesized by several freshwater species of cyanobacteria (i.e. Raphidiopsis raciborskii and Anabaena flos-aquae) and are some of the most common cyanotoxins in aquatic reservoirs. The harmful properties of CYN are wide and primarily include cytotoxicity. To date, several analogs and decomposition products of CYN have been described, which can potentially increase its toxic effects in living organisms. The mode of action of ANTX-a is different than that observed after CYN exposure and involves structures in the nervous system. One of the most frequent situations in which cyanotoxins are introduced into the human body is by skin contact with contaminated water, i.e., during water sports, fishing or agriculture. Unfortunately, to date, knowledge on the influence of CYN, its decomposition products, and ANTX-a on human skin is limited. In this paper, we investigated the impact of CYN, its decomposition products, and ANTX-a on the proliferation of human keratinocytes, which provide a protective barrier on the skin. Moreover, we described the cytotoxic effects developed in the selected cell type and estimated the ability of the keratinocytes to migrate under the influence of the studied cyanotoxins. The obtained results suggest that CYN and its decomposition products at concentrations corresponding to that determined for CYN in nature (1 µg·mL-1) are strong inhibitors of keratinocyte proliferation (70% inhibition within 24 h for pure CYN). The cytotoxic effects of CYN and the CYN decomposition products on keratinocytes was also significant, and the pure toxin (1 µg·mL-1) was estimated to be 35% after 24 h of exposure. Similarly, harmful effects caused by CYN and its byproducts were observed during keratinocyte migration, and the initial form of the toxin (1 µg·mL-1) showed 40% inhibition within 16 h. Different results were obtained for ANTX-a. The toxic effects of this compound on human keratinocytes estimated by the applied tests was observed only at the highest tested concentration (10 µg·mL-1) and after a long period of exposure. The results presented in this paper are, to the best of our knowledge, the first description of the influence of CYN, CYN decomposition products, and ANTX-a on human epidermal cells. Clearly, CYN and its decomposition products are serious threats not only when acting on internal organs but also during the skin contact stage. Further studies on cyanotoxins should focus on the determination of their decomposition products and ecotoxicology in natural aquatic environments.

Mycosporine-Like Amino Acids: Potential Health and Beauty Ingredients.

Human skin is constantly exposed to damaging ultraviolet radiation (UVR), which induces a number of acute and chronic disorders. To reduce the risk of UV-induced skin injury, people apply an additional external protection in the form of cosmetic products containing sunscreens. Nowadays, because of the use of some chemical filters raises a lot of controversies, research focuses on exploring novel, fully safe and highly efficient natural UV-absorbing compounds that could be used as active ingredients in sun care products. A promising alternative is the application of multifunctional mycosporine-like amino acids (MAAs), which can effectively compete with commercially available filters. Here, we outline a complete characterization of these compounds and discuss their enormous biotechnological potential with special emphasis on their use as sunscreens, activators of cells proliferation, anti-cancer agents, anti-photoaging molecules, stimulators of skin renewal, and functional ingredients of UV-protective biomaterials.

Determination of the toxicity of the freshwater cyanobacterium Woronichinia naegeliana (Unger) Elenkin.

Cyanobacterial blooms are undesirable for ecological and health reasons. While Woronichinia naegeliana is a cyanobacterial species that appears frequently in freshwater, information about it is limited. An evaluation of its toxicity was conducted via tests based on the crustaceans Thamnocephalus platyurus and Daphnia pulex. The greatest effect of the aqueous extract obtained from W. naegeliana cells was observed for T. platyurus. The denoted semi-lethal concentration (LC50) after 24 h of exposure was 0.99 mg of dry weight (d.w.) mL-1. A lower toxicity was displayed for D. pulex, although it grew with time. Among the 18 fractions separated from cyanobacterial extract, only one containing the microginin FR3 (MG-FR3) displayed biological activeness against T. platyurus. The remaining products synthesized by W. naegeliana displayed an absence or a low level of toxicity making it impossible to determine the LC50 value. Detailed research revealed that MG-FR3 did not affect the activity of enzymes such as trypsin, chymotrypsin, elastase and thrombin, which indicates another mode of action. The results demonstrated that blooms of W. naegeliana showed toxic activity towards invertebrate zooplankton.

Secondary metabolites of the lichen Hypogymnia physodes (L.) Nyl. and their presence in spruce (Picea abies (L.) H. Karst.) bark.

Lichen species typically have a characteristic profile of secondary metabolites. Dense populations of Hypogymnia physodes growing frequently as epiphytes on tree branches have harmful effects on the host, likely due to their secondary compounds, which were undetected in tree tissues until now. The aim of the present study was to re-characterise the suite of secondary metabolites of H. physodes thalli and to estimate their translocation into spruce (Picea abies) bark. Thallus and bark extracts were compared using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The compounds were identified based on their UV, MS and MS/MS spectra as well as retention factors of their TLC analysis. In addition to the previously described secondary metabolites (protocetraric, physodalic, 3-hydroxyphysodic, physodic, and 2'-O-methylphysodic acids, atranorin and chloroatranorin) of H. physodes, further three were identified in its thalli: conphysodalic, 4-O-methylphysodic and α-alectoronic acids. Fragmentation patterns from the negative ionisation of each compound were proposed, some of which were described for the first time. Among all of the detected lichen substances, a few, e.g., physodalic, 3-hydroxyphysodic, physodic acids and atranorin, were present in the bark of spruce branches that were abundantly colonised by lichen. The newly identified compounds of H. physodes thalli may belong to its constant or accessory secondary metabolites. These compounds may be useful in the chemotaxonomic classification of this species. The presence of some lichen substances in spruce bark confirmed their ability to penetrate host tissues. These data suggest that H. physodes compounds may cause long-term effects on spruces in nature.

Microcystins and anatoxin-a in Arctic biocrust cyanobacterial communities.

In the polar regions cyanobacteria are an important element of plant communities and represent the dominant group of primary producers. They commonly form thick highly diverse biological soil crusts that provide microhabitats for other organisms. Cyanobacteria are also producers of toxic secondary metabolites. In the present study we demonstrated that biocrust-forming cyanobacteria inhabiting the Kaffiøyra Plain, the north-west coast of Spitsbergen, are able to synthesize toxins, especially microcystins (MCs, from 0.123 to 11.058 µg MC-LR per g dry weight, DW) and anatoxin-a (ANTX-a, from 0.322 to 0.633 µg ANTX-a per g DW). To the best of our knowledge, this is the first report on the presence of ANTX-a in the entire polar region. The occurrence of cyanotoxins can exert a long-term impact on organisms co-existing in biocrust communities and can have far-reaching consequences for the entire polar ecosystem.

Phytoremediation of anatoxin-a by aquatic macrophyte Lemna trisulca L.

The neurotoxin anatoxin-a (ANTX-a), one of the most common cyanotoxin, poses a health risk to people and can be lethal to aquatic organisms. This paper presents results on its bioremediation by the aquatic macrophyte Lemna trisulca. We show that the plant is resistant to the harmful impact of toxin and is capable of removing ANTX-a from water. Some of the ANTX-a concentrations which were used in our experiments were much higher than those found in natural conditions. The exposition of L. trisulca to 2.5 µg ANTX-a/mL did not affect its biomass accumulation within 24 d. Significant decreases in biomass content by 21% and 30% were demonstrated in samples cultivated in media containing 12.5 µg ANTX-a/mL after 18 and 24 day of experiment, respectively. One gram of fresh weight (f.w.) of L. trisulca cultured for 14 d in the media containing 50 µg ANTX-a removed 95% of the initial toxin concentration; for media with 250 µg ANTX-a, 86% was removed. In tests of ANTX-a binding stability and degradation we transferred the macrophyte to fresh media without added toxin; within 14 d the content of accumulated ANTX-a in the macrophyte decreased by 76% (from initial 19.3 µg ANTX-a/gf.w.), 71% (from 37.3 µg ANTX-a/g f.w.) and 47% (from 63.7 µg ANTX-a/g f.w.). The quantity of ANTX-a released to media was minimal: from 3.5% to 5.1% of the initial bioaccumulated value. The data show that part of the ANTX-a was degraded. Mass spectra analyses did not indicate transformation of ANTX-a to already known forms. These findings suggest that L. trisulca has much potential as a phytoremediation agent for stabilization of aquatic environments.

EXTRACELLULAR ENZYMES OF THE MICROCYSTIS AERUGINOSA PCC 7813 STRAIN ARE INHIBITED IN THE PRESENCE OF HYDROQUINONE AND PYROGALLOL, ALLELOCHEMICALS PRODUCED BY AQUATIC PLANTS(1).

Several cyanobacterial species have a high potential to dominate in marine environments and freshwater reservoirs, and the ecological and physiological reasons for this phenomenon are not understood comprehensively. In this study, the ability of a Microcystis aeruginosa Kütz. strain to produce free dissolved enzymes was documented. We have observed that this highly toxic strain releases alkaline phosphatase, leucine aminopeptidase, and ß-glucosidase into the ambient environment. Additionally, the inhibitory activity of selected phenols produced by aquatic plants on the activity of these enzymes was analyzed. The investigated compounds, pyrogallol and, to a lesser degree, hydroquinone, decreased the activity of extracellular enzymes produced by M. aeruginosa, with leucine aminopeptidase being the most sensitive to the inhibitors. The noncompetitive character of enzymatic inhibition suggests that the polyphenols produced by aquatic plants are able to influence the activity of different extracellular or membrane-bound enzymes.

Degradation of microcystin-LR by ozone in the presence of Fenton reagent.

In this study the effectiveness and feasibility of the degradation of microcystin-LR (MC-LR) using a combined method ozone/Fenton reagent was investigated. The decomposition of the toxin was determined by the chromatographic technique. The effect of the Fenton reaction on the ozonation was observed at various concentrations of ozone at pH 3.0 and 6.8. A low concentration of ozone (0.01 mg/L) given simultaneously with Fenton reagent was more effective in MC-LR degradation than ozone or Fenton reagent treatment individually regardless of the pH. At higher concentrations of ozone the use together with Fenton reagent at pH 6.8 was less efficient in MC-LR degradation than treatment with ozone alone, whereas at pH 3.0 Fenton process turned out to be more effective. The mixture of ozone and hydrogen peroxide was more striking than using ozone with Fenton reagent regardless of the pH. The complete degradation of MC-LR was achieved using ozone alone at a concentration of 0.10 mg/L, whereas the same result using ozonation with Fenton reagent required a dose of ozone of 0.20 mg/L at pH 6.8. In acidic pH the total removal of MC-LR was achieved using ozone alone at a concentration of 0.20 mg/L and the same result was obtained for the combined method of ozone/Fenton reagent.

The alteration of Microcystis aeruginosa biomass and dissolved microcystin-LR concentration following exposure to plant-producing phenols.

Plant-producing phenols could strongly inhibit the growth of toxic cyanobacteria genera, but the ecological consequences of this action are still unknown. In this work, the activity of selected phenols in relation to the strain Microcystis aeruginosa was investigated. We have found that the mechanism of the growth inhibition could involve both the inhibition of the photosynthetic system and the alkaline phosphatase activity. The excretion of a high amount of microcystin-LR to the environment was observed as a final result of the application of phenols to the cyanobacterial culture. This finding suggests that first of all an algicidal effect of phenols was probably the reason of the cyanobacterial biomass decreasing.

Sensitised decomposition of microcystin-LR using UV radiation.

UV radiation was applied to degrade cyanobacterial hepatotoxin, microcystin-LR in the presence of phycocyanin as a model natural sensitiser. The concentrations of both the toxin and the pigment used in the experiments were higher by several orders of magnitude than found in the environment. The photoreaction parameters were optimised. The process was found to be of limited use for water treatment due to its low efficacy. Additionally, pronounced UV-induced bleaching of the pigment significantly reducing the photoreaction rates of the toxin was observed for the highest UV radiation intensities applied.

THE EFFECT OF ABSCISIC ACID AND FUSICOCCIN ON MALIC ACID CONCENTRATION IN PULVINI OF PHASEOLUS COCCINEUS L.

Fusicoccin and abscisic acid can modify the movements of some nyctinastic plants. This paper reports the effects of these two compounds on the concentrations of malic acid in pulvini of Phaseolus coccineus L. in the transition period of two phases of the circadian rhythm. The concentrations of malic acid in the two parts of the pulvinus, extensor and flexor, depend upon the phase of leaf movements. Abscisic acid and fusicoccin significantly affected the content of malic acid in leaves compared with the control. In the two phases of the diurnal cycle investigated, abscisic acid markedly reduced the production of malic acid in the flexor and extensor regions of the pulvinus, and treatment with fusicoccin produced small but distinct increases in the concentration of malic acid in the same regions. The results are discussed with particular reference to the effect of abscisic acid and fusicoccin on the activity of the H+ pump associated with the control of turgor in cells.