The effect of acute high light and low temperature stresses on the ascorbate–glutathione cycle and superoxide dismutase activity in two Dunaliella salina strains

Dunaliella species accumulate carotenoids and their role in protection against photooxidative stress has been investigated extensively. By contrast, the role of other antioxidants in this alga, has received less attention. Therefore, the components of the ascorbate–glutathione cycle, along with superoxide dismutase (E.C. 1.15.1.1) and peroxidase (E.C. 1.11.1.11) activity were compared in two strains of Dunaliella salina. Strain IR‐1 had two‐fold higher chlorophyll and β‐carotene concentration than Gh‐U. IR‐1 had around four‐fold higher superoxide dismutase, ascorbate peroxidase and pyrogallol peroxidase activities than Gh‐U on a protein basis. Ascorbate and glutathione concentrations and redox state did not differ between strains and there was little difference in the activity of ascorbate–glutathione cycle enzymes (monodehydroascorbate reductase [E.C. 1.6.5.4], dehydroascorbate reductase [E.C. 1.8.5.1] and glutathione reductase [E.C. 1.8.1.7]). The response of these antioxidants to high light and low temperature was assessed by transferring cells from normal growth conditions (28°C, photon flux density of 100 μmol m^?2 s^?1)to 28°C/1200 μmol m^?2 s^?1; 13°C/100 μmol m^?2 s^?1; 13°C/1200 μmol m^?2 s^?1 and 28°C/100 μmol m^?2 s^?1 for 24 h. Low temperature and combined high light‐low temperature decreased chlorophyll and β‐carotene in both strains indicating that these treatments cause photooxidative stress. High light, low temperature and combined high light‐low temperature treatments increased the total ascorbate pool by 10–50% and the total glutathione pool by 20–100% with no consistent effect on their redox state. Activities of ascorbate–glutathione cycle enzymes were not greatly affected but all the treatments increased superoxide dismutase activity. It is concluded that D. salina can partially adjust to photooxidative conditions by increasing superoxide dismutase activity, ascorbate and glutathione.     

High Levels of Ascorbic Acid, Not Glutathione, in the CNS of Anoxia-Tolerant Reptiles Contrasted with Levels in Anoxia-Intolerant Species

Abstract: Ascorbic acid and glutathione (GSH) are antioxidants and free radical scavengers that provide the first line of defense against oxidative damage in the CNS. Using HPLC with electrochemical detection, we determined tissue contents of these antioxidants in brain and spinal cord in species with varying abilities to tolerate anoxia, including anoxia-tolerant pond and box turtles, moderately tolerant garter snakes, anoxia-intolerant clawed frogs (Xenopus laevis), and intolerant Long-Evans hooded rats. These data were compared with ascorbate and GSH levels in selected regions of guinea pig CNS, human cortex, and values from the literature. Ascorbate levels in turtles were typically 100% higher than those in rat. Cortex, olfactory bulb, and dorsal ventricular ridge had the highest content in turtle, 5–6 µmol g^?1 of tissue wet weight, which was twice that in rat cortex (2.82 ± 0.05 µmol g^?1) and threefold greater than in guinea pig cortex (1.71 ± 0.03 µmol g^?1). Regionally distinct levels (2–4 µmol g^?1) were found in turtle cerebellum, optic lobe, brainstem, and spinal cord, with a decreasing anterior-to-posterior gradient. Ascorbate was lowest in white matter (optic nerve) in each species. Snake cortex and brainstem had significantly higher ascorbate levels than in rat or guinea pig, although other regions had comparable or lower levels. Frog ascorbate was generally in an intermediate range between that in rat and guinea pig. In contrast to ascorbate, GSH levels in anoxia-tolerant turtles, 2–3 µmol g^?1 of tissue wet weight, were similar to those in mammalian or amphibian brain, with no consistent pattern associated with anoxia tolerance. GSH levels in pond turtle CNS were significantly higher (by 10–20%) than in rat for several regions but were generally lower than in guinea pig or frog. GSH in box turtle and snake CNS were the same or lower than in rat or guinea pig. The distribution GSH in the CNS also had a decreasing anterior-to-posterior gradient but with less variability than ascorbate; levels were similar in optic nerve, brainstem, and spinal cord. The paradoxically high levels of ascorbate in turtle brain, which has a lower rate of oxidative metabolism than mammalian, suggest that ascorbate is an essential cerebral antioxidant. High levels may have evolved to protect cells from oxidative damage when aerobic metabolism resumes after a hypoxic dive.     

Overexpression of bacterial catalase in tomato leaf chloroplasts enhances photo-oxidative stress tolerance

The Escherichia coli gene katE, which is driven by the promoter of the Rubisco small subunit gene of tomato, rbcS3C, was introduced into a tomato (Lycopersicon esculentum Mill.) by Agrobacterium tumefaciens‐mediated transformation. Catalase activity in progeny from transgenic plants was approximately three‐fold higher than that in wild‐type plants. Leaf discs from transgenic plants remained green at 24 h after treatment with 1 µm paraquat under moderate light intensity, whereas leaf discs from wild‐type plants showed severe bleaching after the same treatment. Moreover, ion leakage from transgenic leaf discs was significantly less than that from wild‐type leaf discs at 24 h after treatment with 1 µm paraquat and 10 mm H₂O₂, respectively, under moderate light intensity. To evaluate the efficiency of the E. coli catalase to protect the whole transgenic plant from the oxidative stress, transgenic and wild‐type plants were sprayed with 100 µm paraquat and exposed to high light illumination (800 µmol m^?2 s^?1). After 24 h, the leaves of the transgenic plants were less damaged than the leaves of the wild‐type plants. The catalase activity and the photosynthesis activity (indicated by the F_v/F_m ratio) were less affected by paraquat treatment in leaves of transgenic plants, whereas the activities of the chloroplastic ascorbate peroxidase isoenzymes and the ascorbate content decreased in both lines. In addition, the transgenic plants showed increased tolerance to the oxidative damage (decrease of the CO₂ fixation and photosystem II activity and increase of the lipid peroxidation) caused by drought stress or chilling stress (4 °C) under high light intensity (1000 µmol m^?2 s^?1). These results indicate that the expression of the catalase in chloroplasts has a positive effect on the protection of the transgenic plants from the photo‐oxidative stress invoked by paraquat treatment, drought stress and chilling stress.     

Photosynthetic and biochemical characterization of in vitro-derived African violet (Saintpaulia ionantha H. Wendl) plants to ex vitro conditions

African violet (Saintpaulia ionantha H. Wendl) is one of the most easily and commonly tissue-cultured ornamental plants. Despite this, there are limited reports on photosynthetic capacity and its impact on the plant quality during acclimatization. Various growth, photosynthetic and biochemical parameters and activities of antioxidant enzymes and dehydrins of micropropagated plants were assessed under three light intensities (35, 70, and 100 µmol m^?2 s^?1 photosynthetic photon flux density – PPFD). Fresh and dry plant biomass, plant height, and leaf area were optimal with high irradiance (70–100 µmol m^?2 s^?1 PPFD). Chlorophyll and carotenoid contents and net photosynthesis were optimal in plants grown under 70 µmol m^?2 s^?1 PPFD. Stomatal resistance, malondialdehyde content, and F_v/F_m values were highest at low light irradiance (35 µmol m^?2 s^?1 PPFD). The activities of three antioxidant enzymes, superoxide dismutase, catalase, and glutathione peroxidase, increased as light irradiance increased, signaling that high light irradiance was an abiotic stress. The accumulation of 55, 33, and 25 kDa dehydrins was observed with all light treatments although the expression levels were highest at 35 µmol m^?2 s^?1 PPFD. Irradiance at 70 µmol m^?2 s^?1 PPFD was suitable for the acclimatization of African violet plants. Both low and high irradiance levels (35 and 100 µmol m^?2 s^?1 PPFD) induced the accumulation of antioxidants and dehydrins in plants which reveals enhanced stress levels and measures to counter it.     

Changes in adenosine deaminase activity in ageing cultured human cells and the role of zinc

The level of adenosine deaminase (ADA; EC 3.5.4.4) was estimated at different passages in six confluent fibroblast cultures established from forearm skin biopsies of healthy adult normal volunteers. After determination of the zinc concentration in standard growth medium, ADA activity was estimated at different passages of subculture in media with different zinc concentrations. The results indicated that the specific activity of ADA in control confluent skin fibroblast cultures (passage 2) cultivated in standard growth medium containing 15.4 microM zinc (similar to that present in normal human plasma) was equal to 226.6+/-19.64 micromol min(-1) mg(-1) protein. The results showed that there were no significant changes in ADA specific activity in any of the control cultures as the zinc concentration of the medium was increased. To characterize the passage of subculture at which fibroblasts enter the ageing phase, three marker enzymes were assayed namely, phosphofructokinase, lactate dehydrogenase and glycogen phosphorylase. The result showed that the cells enter the ageing phase at passage 20 and beyond. Further investigation showed that ADA activity of serially subcultured confluent cultures cultivated in standard growth medium significantly dropped at passages 20, 25 and 30. ADA activity however was not significantly altered in cells at passage 2, 10 and 15 cultivated in standard growth medium and in the presence of higher zinc levels (23.1, 34.6, 53.8 and 73.1 microM). Furthermore there was significant lowering of ADA activities in cells at passages 20, 25 and 30 when cells were cultured in the presence of 15.4, 23.1 and 34.6 microM zinc. Such lowered activities of ADA were restored to normal when the cells were cultured in the presence of higher zinc concentration equal to 53.8 and 73.1 microM. From the results we concluded that it is possible to restore ADA activity in aged skin fibroblasts to normal levels by raising the zinc concentration in the culture medium to four or five times the control normal plasma zinc level.     

Rapid, reversible alterations in spinach thylakoid appression upon changes in light intensity

Changes in light quantity and quality cause structural changes within the thylakoid membrane; long‐term responses have been described for so‐called ‘sun’ and ‘shade’ leaves. Many leaves, however, experience changes in irradiance on a time scale of minutes due to self‐shading and sun flecks. In this study, mature, attached spinach leaves were grown at 300 µmol photons m^?2 s^?1 then rapidly switched to a different light treatment. The treatment irradiances were 10, 800 or 1500 µmol m^?2 s^?1 for 10 min, or 10 or 20 min of self‐shading (about 10 µmol m^?2 s^?1). Image analysis of transmission electron micrographs revealed that a 10 min switch to a lower light intensity increased grana size and number per chloroplast profile by 10–20%. Returning the leaves to 300 µmol m^?2 s^?1 for 10 min reversed the phenomenon. Chlorophyll fluorescence measurements of detached, intact leaves at 77 K were suggestive of a transition from state 2 to state 1 upon shading. Diurnal ultrastructural measurements of granal size and number did not reveal a significant net change in ultrastructure over the time scale of hours. It is concluded that spinach chloroplasts can alter the degree of thylakoid appression in response to irradiance changes on a time scale of minutes. These ultrastructural responses are caused by biochemical and biophysical adjustments within the thylakoid membrane that serve to maximize photosynthesis and minimize photo‐inhibition under rapidly fluctuating light environments.     

Effects of salinity,light intensity and sediment on growth,pigments, agar production and reproduction in Gracilaria tenuistipitata from Songkhla Lagoon in Thailand

The effects of salinity, light intensity and sediment on Gracilaria tenuistipitata C.F. Chang & B.M. Xia on growth, pigments, agar production, and net photosynthesis rate were examined in the laboratory under varying conditions of salinity (0, 25 and 33 psu), light intensity (150, 400, 700 and 1000 µmol photons m^?2 s^?1) and sediment (0, 0.67 and 2.28 mg L^?1). These conditions simulated field conditions, to gain some understanding of the best conditions for cultivation of G. tenuistipitata. The highest growth rate was at 25 psu, 700 µmol photons m^?2 s^?1 with no sediments, that provided a 6.7% increase in weight gain. The highest agar production (24.8 ± 3.0 %DW) was at 25 psu, 150–400 µmol photons m^?2 s^?1 and no sediment. The highest pigment contents were phycoerythrin (0.8 ± 0.5 mg g^?1FW) and phycocyanin (0.34 ± 0.05 mg g^?1 FW) produced in low light conditions, at 150 µmol photons m^?2 s^?1. The highest photosynthesis rate was 161.3 ± 32.7 mg O₂ g^?1 DW h^?1 in 25 psu, 400 µmol photons m^?2 s^?1 without sediment in the short period of cultivation, (3 days) and 60.3 ± 6.7 mg O₂ g^?1 DW h^?1 in 25 psu, 700 µmol photons m^?2 s^?1 without sediment in the long period of cultivation (20 days). The results indicated that salinity was the most crucial factor affecting G. tenuistipitata growth and production. This would help to promote the cultivation of Gracilaria cultivation back into the lagoon using these now determined baseline conditions. Extrapolation of the results from the laboratory study to field conditions indicated that it was possible to obtain two crops of Gracilaria a year in the lagoon, with good yields of agar, from mid‐January to the end of April (dry season), and from mid‐July to the end of September (first rainy season) when provided sediment was restricted.     

Green synthesis of fluorescence carbon nanoparticles from yum and application in sensitive and selective detection of ATP

Fluorescent carbon nanoparticles (CPs), a fascinating class of recently discovered nanocarbons, have been widely known as some of the most promising sensing probes in biological or chemical analysis. In this study, we demonstrate a green synthetic methodology for generating water‐soluble CPs with a quantum yield of approximately 24% via a simple heating process using yum mucilage as a carbon source. The prepared carbon nanoparticles with an ~10 nm size possessed excellent fluorescence properties, and the fluorescence of the CPs was strongly quenched by Fe³⁺, and recovered by adenosine triphosphate (ATP), thus, an ‘off’ and ‘on’ system can be easily established. This ‘CPs‐Fe³⁺‐ATP’ strategy was sensitive and selective at detecting ATP with the linear range of 0.5 µmol L^?1 to 50 µmol L^?1 and with a detection limit of 0.48 µmol L^?1. Copyright © 2015 John Wiley & Sons, Ltd.     

Is digestive capacity limiting growth at low temperatures in roach?

In roach Rutilus rutilus growth ceases below a temperature threshold of 12° C. This cessation of growth is accompanied by a reduction in feeding. Do roach decrease feeding in the cold because of reduced energy demand, caused by the decelerating effect of low temperature on metabolism and growth, or is feeding directly limited by low temperatures, leading to reduced growth rates? It was found that at low temperatures the intake and digestion of food may be limited by reduced activities of digestive enzymes. Trypsin, amylase and γ‐glutamyl transferase showed a negative compensation with respect to temperature, resulting in very low activities at acclimation temperatures of ≤12° C. Trypsin activity, falling from 400·5 ± 131·2 U g^?1 fresh mass of the gut at 27° C to 12·5 U g^?1 fresh mass at 4° C, displayed the strongest linear correlation with growth rates, suggesting that trypsin activities may set a limit to growth in the low temperature range. If protein digestion is limiting at low temperatures, this should be reflected in reduced concentrations of amino acid in the white muscle. The size of the total amino acid pool was not affected by temperature acclimation and ranged between 19·2 ± 6·2 and 25·2 ± 3·6 µmol g^?1 fresh mass of the white muscle. A decrease, however, was found of several amino acids, mainly of threonine and glutamine, in the low temperature range. Low concentrations of the essential amino acid threonine (0·14 ± 0·03 µmol g^?1 fresh mass at 12° C and 0·12 ± 0·05 µmol g^?1 fresh mass at 4° C) were probably due to nutritional or digestional limitations and may therefore have resulted from reduced trypsin activity in the cold. The non‐essential amino acid glutamine, however, can be endogenously synthesized and its low level observed at 4° C (0·16 ± 0·09 µmol g^?1 fresh mass) was not necessarily a result of low trypsin activities. It is more likely that low temperatures impair glutamine synthesis. The possibility that glutamine concentrations may be down regulated under conditions when anabolic processes are not advantageous is discussed.     

Photon irradiance required to support optimal growth and interrelations between irradiance and pigment composition in the green alga Haematococcus pluvialis

The aim of the work was to find the optimal photon irradiance for the growth of green cells of Haematococcus pluvialis and to study the interrelations between changes in photochemical parameters and pigment composition in cells exposed to photon irradiances between 50 and 600?µmol?m^?2?s^?1 and a light:dark cycle of 12:12?h. Productivity of cultures increased with irradiance. However, the rate of increase was higher in the range 50–200?µmol?^?2?s^?1. The carotenoid content increased with increasing irradiance, while the chlorophyll content decreased. The maximum quantum yield of PSII (F_v/F_m) gradually declined from 0.76 at the lowest irradiance of 50?µmol?^?2?s^?1 to 0.66 at 600?µmol?^?2?s^?1. Photosynthetic activity showed a drop at the end of the light period, but recovered fully during the following dark phase. A steep increase in non-photochemical quenching was observed when cultures were grown at irradiances above 200?µmol?^?2?s^?1. A sharp increase in the content of secondary carotenoids also occurred above 200?µmol?m^?2?s^?1. According to our results, with H. pluvialis green cells grown in a 5-cm light path device, 200?µmol?^?2?s^?1 was optimal for growth, and represented a threshold above which important changes in both photochemical parameters and pigment composition occurred.     

'The effect of micronutrients on superoxide dismutase in senescent fibroblasts'

The specific activities of zinc/copper (Zn/Cu)-superoxide dismutase (SOD-1) and manganese (Mn)-superoxide dismutase (SOD-2) were assayed in young passage 5 fibroblasts and in serially subcultured cells that were characterized as senescent at passages 15-35. SOD-1 and SOD-2 activities did not significantly change in senescent and young cells cultured in either routine medium [minimum essential medium 1 (MEM1)], or in Zn, Cu and Mn supplemented medium (MEM2) containing normal human plasma levels of the cations. SOD-1 and SOD-2 activities, however, underwent parallel progressive significant activity increases in senescent passage 20 and 25 cells, which peaked in value in passage 30 and 35 cells subcultured in supplemented medium (MEM3) containing triple human plasma levels of the cations. Concurrently, superoxide radical generation rates underwent progressive significant increases in senescent passage 15-25 cells, which peaked in value in passage 30 and 35 cells subcultured in MEM1 or MEM2. These rates, however, were significantly lowered in senescent cells subcultured in MEM3. We infer that it was only possible to significantly stimulate SOD-1 and SOD-2 activities in senescent MEM3 cultured cells enabling them to combat oxidative stress.     

Continuous 3D Titanium Nitride Nanoshell Structure for Solar‐Driven Unbiased Biocatalytic CO2 Reduction

Z‐scheme‐inspired tandem photoelectrochemical (PEC) cells have received attention as a sustainable platform for solar‐driven CO₂ reduction. Here, continuously 3D‐structured, electrically conductive titanium nitride nanoshells (3D TiN) for biocatalytic CO₂‐to‐formate conversion in a bias‐free tandem PEC system are reported. The 3D TiN exhibits a periodically porous network with high porosity (92.1%) and conductivity (6.72 × 10⁴ S m^?1), which allows for high enzyme loading and direct electron transfer (DET) to the immobilized enzyme. It is found that the W‐containing formate dehydrogenase from Clostridium ljungdahlii (ClFDH) on the 3D TiN nanoshell is electrically activated through DET for CO₂ reduction. At a low overpotential of 40 mV, the 3D TiN‐ClFDH stably converts CO₂ to formate at a rate of 0.34 µmol h^?1 cm^?2 and a faradaic efficiency (FE) of 93.5%. Compared to a flat TiN‐ClFDH, the 3D TiN‐ClFDH shows a 58 times higher formate production rate (1.74 µmol h^?1 cm^?2) at 240 mV of overpotential. Lastly, a bias‐free biocatalytic tandem PEC cell that converted CO₂ to formate at an average rate of 0.78 µmol h^?1 and an FE of 77.3% only using solar energy and water is successfully assembled.     

Phytochrome modulation of blue-light-induced phototropism

Red light enhances hypocotyl phototropism toward unilateral blue light through a phytochrome‐mediated response. This study demonstrates how the phytochromes modulate blue‐light‐induced phototropism in the absence of a red light pre‐treatment. It was found that phytochromes A, B, and D have conditionally overlapping functions in the promotion of blue‐light‐induced phototropism. Under very low blue light intensities (0.01 µmol m^?2 s^?1) phyA activity is necessary for the progression of a normal phototropic response, whereas above 1.0 µmol m^?1 s^?2 phyB and phyD have functional redundancy with phyA to promote phototropism. PhyA also contributes to attenuation of phototropism under high fluence rates of unilateral blue light, which was previously shown to be dependent on the phototropins and cryptochromes. From these results, it appears that phytochromes are required to develop a robust phototropic response under low fluence rates, whereas under high irradiances where phototropism may be less important, phyA suppresses phototropism.     

The biotechnological potential of subtilisin‐like fibrinolytic enzyme from a newly isolated Lactobacillus plantarum KSK‐II in blood destaining and antimicrobials

An antimicrobial oxidative‐ and SDS‐stable fibrinolytic alkaline protease designated as KSK‐II was produced by Lactobacillus plantarum KSK‐II isolated from kishk, a traditional Egyptian food. Maximum enzyme productivity was obtained in medium containing 1% lactose and 0.5% soybean flour as carbon and nitrogen sources, respectively. Purification of enzyme increased its specific activity to 1,140‐fold with a recovery of 33% and molecular weight of 43.6 kDa. Enzyme activity was totally lost in the presence of ethylenediaminetetraacetic acid and was restored after addition of Fe²⁺ suggesting that KSK‐II is a metalloprotease and Fe²⁺ acts as cofactor. Enzyme hydrolyzed not only the natural proteins but also synthetic substrates, particularly Suc‐Ala‐Ala‐Pro‐Phe‐pNA. KSK‐II can hydrolyze the Lys‐X easier than Arg‐X; thus, it was considered as a subtilisin‐family protease. Its apparent K_m, V_max, and K_cat were 0.41 mM, 6.4 µmol mg^?1 min^?1, and 28.0 s^?1, respectively. KSK‐II is industrially important from the perspectives of its maximal activity at 50°C (stable up to 70°C), ability to function at alkaline pH (10.0), stability at broad pH ranges (7.5–12.0) in addition to its stability toward SDS, H₂O₂, organic solvents, and detergents. We emphasize for the first time the potential of fibrinolytic activity for alkaline proteases used in detergents especially in blood destaining. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 31:316–324, 2015     

Net grassland carbon flux over a subambient to superambient CO2 gradient

Increasing atmospheric CO₂ concentrations may have a profound effect on the structure and function of plant communities. A previously grazed, central Texas grassland was exposed to a 200‐µmol mol^?1 to 550 µmol mol^?1 CO₂ gradient from March to mid‐December in 1998 and 1999 using two, 60‐m long, polyethylene‐ covered chambers built directly onto the site. One chamber was operated at subambient CO₂ concentrations (200–360 µmol mol^?1 daytime) and the other was regulated at superambient concentrations (360–550 µmol mol^?1). Continuous CO₂ gradients were maintained in each chamber by photosynthesis during the day and respiration at night. Net ecosystem CO₂ flux and end‐of‐year biomass were measured in each of 10, 5‐m long sections in each chamber. Net CO₂ fluxes were maximal in late May (c. day 150) in 1998 and in late August in 1999 (c. day 240). In both years, fluxes were near zero and similar in both chambers at the beginning and end of the growing season. Average daily CO₂ flux in 1998 was 13 g CO₂ m^?2 day^?1 in the subambient chamber and 20 g CO₂ m^?2 day^?1 in the superambient chamber; comparable averages were 15 and 26 g CO₂ m^?2 day^?1 in 1999. Flux was positively and linearly correlated with end‐of‐year above‐ground biomass but flux was not linearly correlated with CO₂ concentration; a finding likely to be explained by inherent differences in vegetation. Because C₃ plants were the dominant functional group, we adjusted average daily flux in each section by dividing the flux by the average percentage C₃ cover. Adjusted fluxes were better correlated with CO₂ concentration, although scatter remained. Our results indicate that after accounting for vegetation differences, CO₂ flux increased linearly with CO₂ concentration. This trend was more evident at subambient than superambient CO₂ concentrations.     

High-temperature inhibition of photosynthesis is greater under sunflecks than uniform irradiance in a tropical rain forest tree seedling

The survival of dipterocarp seedlings in the understorey of south‐east Asian rain forests is limited by their ability to maintain a positive carbon balance. Photosynthesis during sunflecks is an important component of carbon gain. Field measurements demonstrated that Shorea leprosula seedlings in a rain forest understorey received a high proportion of daily photon flux density at temperatures supra‐optimal for photosynthesis (72% at ≥30 °C, 14% at ≥35 °C). To investigate the effect of high temperatures on photosynthesis during sunflecks, gas exchange and chlorophyll fluorescence measurements were made on seedlings grown in controlled environment conditions either, under uniform, saturating irradiance (approximately 539 µmol m^?2 s^?1) or, shade/fleck sequences (approximately 30 µmol m^?2 s^?1/approximately 525 µmol m^?2 s^?1) at two temperatures, 28 or 38 °C. The rate of light‐saturated photosynthesis, under uniform irradiance, was inhibited by 40% at 38 °C compared with 28 °C. However, during the shade/fleck sequence, photosynthesis was inhibited by 59% at 38 °C compared with 28 °C. The greater inhibition of photosynthesis during the shade/fleck sequence, when compared with uniform irradiance, was driven by the lower efficiency of dynamic photosynthesis combined with lower steady‐state rates of photosynthesis. These results suggest that, contrary to current dogma, sunfleck activity may not always result in significant carbon gain. This has important consequences for seedling regeneration processes in tropical forests as well as for leaves in other canopy positions where sunflecks make an important contribution to total photon flux density.     

Microflow injection potassium bioassay based on G‐quadruplex DNAzyme‐enhanced chemiluminescence

By taking advantage of microflow injection chemiluminescence analysis, we developed a distinctive microfluidic bioassay method based on G‐Quadruplex DNAzyme‐enhanced chemiluminescence for the determination of K⁺ in human serum. AGRO100, the G‐rich oligonucleotide with high hemin binding affinity was primarily selected as a K⁺ recognition element. In the presence of K⁺, AGRO100 folded into G‐quadruplex and bound hemin to form DNAzyme, which catalyzed the oxidation of luminol by H₂O₂ to produce chemiluminescence. The intensity of chemiluminescence increased with the K⁺ concentration. In the study, the DNAzyme showed both long‐term stability and high catalytic activity; other common cations at their physiological concentration did not cause notable interference. With only 6.7 × 10^?13 mol of AGRO100 consumption per sample, a linear response of K⁺ ranged from 1 to 300 µmol/L, the concentration detection limit 0.69 µmol/L (S/N = 3) and the absolute detection limit 1.38 × 10^?12 mol were obtained. The precision of 10 replicate measurements of 60 µmol/L K⁺ was found to be 1.72% (relative standard deviation). The accuracy of the method was demonstrated by analyzing real human serum samples. Copyright © 2014 John Wiley & Sons, Ltd.     

Soil-surface carbon dioxide efflux and microbial biomass in relation to tree density 13 years after a stand replacing fire in a lodgepole pine ecosystem

The effects of fire on soil‐surface carbon dioxide (CO₂) efflux, F_S, and microbial biomass carbon, C_mic, were studied in a wildland setting by examining 13‐year‐old postfire stands of lodgepole pine differing in tree density (< 500 to > 500 000 trees ha^?1) in Yellowstone National Park (YNP). In addition, young stands were compared to mature lodgepole pine stands (～110‐year‐old) in order to estimate ecosystem recovery 13 years after a stand replacing fire. Growing season F_S increased with tree density in young stands (1.0 µmol CO₂ m^?2 s^?1 in low‐density stands, 1.8 µmol CO₂ m^?2 s^?1 in moderate‐density stands and 2.1 µmol CO₂ m^?2 s^?1 in high‐density stands) and with stand age (2.7 µmol CO₂ m^?2 s^?1 in mature stands). Microbial biomass carbon in young stands did not differ with tree density and ranged from 0.2 to 0.5 mg C g^?1 dry soil over the growing season; C_mic was significantly greater in mature stands (0.5–0.8 mg C g^?1 dry soil). Soil‐surface CO₂ efflux in young stands was correlated with biotic variables (above‐ground, below‐ground and microbial biomass), but not with abiotic variables (litter and mineral soil C and N content, bulk density and soil texture). Microbial biomass carbon was correlated with below‐ground plant biomass and not with soil carbon and nitrogen, indicating that plant activity controls not only root respiration, but C_mic pools and overall F_S rates as well. These findings support recent studies that have demonstrated the prevailing importance of plants in controlling rates of F_S and suggest that decomposition of older, recalcitrant soil C pools in this ecosystem is relatively unimportant 13 years after a stand replacing fire. Our results also indicate that realistic predictions and modeling of terrestrial C cycling must account for the variability in tree density and stand age that exists across the landscape as a result of natural disturbances.     

Dynamic changes of canopy-scale mesophyll conductance to CO₂ diffusion of sunflower as affected by CO₂ concentration and abscisic acid

Leaf‐level measurements have shown that mesophyll conductance (g_m) can vary rapidly in response to CO₂ and other environmental factors, but similar studies at the canopy‐scale are missing. Here, we report the effect of short‐term variation of CO₂ concentration on canopy‐scale g_m and other CO₂ exchange parameters of sunflower (Helianthus annuus L.) stands in the presence and absence of abscisic acid (ABA) in their nutrient solution. g_m was estimated from gas exchange and on‐line carbon isotope discrimination (Δ_obs) in a ¹³CO₂/¹²CO₂ gas exchange mesocosm. The isotopic contribution of (photo)respiration to stand‐scale Δ_obs was determined with the experimental approach of Tcherkez et al. Without ABA, short‐term exposures to different CO₂ concentrations (C_a 100 to 900 µmol mol^?1) had little effect on canopy‐scale g_m. But, addition of ABA strongly altered the CO₂‐response: g_m was high (approx. 0.5 mol CO₂ m^?2 s^?1) at C_a < 200 µmol mol^?1 and decreased to <0.1 mol CO₂ m^?2 s^?1 at C_a >400 µmol mol^?1. In the absence of ABA, the contribution of (photo)respiration to stand‐scale Δ_obs was high at low C_a (7.2‰) and decreased to <2‰ at C_a > 400 µmol mol^?1. Treatment with ABA halved this effect at all C_a.     

Durum wheat seedling responses to simultaneous high light and salinity involve a fine reconfiguration of amino acids and carbohydrate metabolism

Durum wheat plants are extremely sensitive to drought and salinity during seedling and early development stages. Their responses to stresses have been extensively studied to provide new metabolic targets and improving the tolerance to adverse environments. Most of these studies have been performed in growth chambers under low light [300–350 µmol m^?2 s^?1 photosynthetically active radiation (PAR), LL]. However, in nature plants have to face frequent fluctuations of light intensities that often exceed their photosynthetic capacity (900–2000 µmol m^?2 s^?1). In this study we investigated the physiological and metabolic changes potentially involved in osmotic adjustment and antioxidant defense in durum wheat seedlings under high light (HL) and salinity. The combined application of the two stresses decreased the water potential and stomatal conductance without reducing the photosynthetic efficiency of the plants. Glycine betaine (GB) synthesis was inhibited, proline and glutamate content decreased, while γ‐aminobutyric acid (GABA), amides and minor amino acids increased. The expression level and enzymatic activities of Δ1‐pyrroline‐5‐carboxylate synthetase, asparagine synthetase and glutamate decarboxylase, as well as other enzymatic activities of nitrogen and carbon metabolism, were analyzed. Antioxidant enzymes and metabolites were also considered. The results showed that the complex interplay seen in durum wheat plants under salinity at LL was simplified: GB and antioxidants did not play a main role. On the contrary, the fine tuning of few specific primary metabolites (GABA, amides, minor amino acids and hexoses) remodeled metabolism and defense processes, playing a key role in the response to simultaneous stresses.