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The enzyme superoxide dismutase (SOD) holds a key position in the microalgal antioxidant network. The present research focused on oxidative stress responses in the Antarctic diatom Chaetoceros brevis F. Schutt during transition to excess (including ultra- violet radiation (UVR)) and limiting irradiance con- ditions. Over a 4 d period, cellular responses of thiobarbituric acid reactive substances (TBARS, a general oxidative stress indicator), SOD activity, photosynthetic and xanthophyll cycle pigments, PSII efficiency, and growth were determined. In addition, oxidative responses were measured during a daily cycle. Changing irradiance conditions significantly affected growth rates of C. brevis. PSII efficiency decreased significantly during periodic excess irradi- ance and increased under low irradiance conditions. Transition to excess irradiance increased the ratio of xanthophyll to light-harvesting pigments, whereas the opposite was observed for cultures transferred to low irradiance. This acclimation process was com- pleted after 2 d in the new irradiance environment. SOD activity increased significantly after the first day regardless of the new irradiance environment but returned to preexposure values on the fourth day. We hypothesize that SOD activity may be tem- porarily elevated in C. brevis after irradiance shifts, thereby reducing oxidative stress when photoaccli- mation is in progress.

The synergistic effects of iron limitation and irradiance dynamics on growth, photosynthesis, antioxidant activity and excessive PAR (400-700 nm) and UV (280-400 nm) sensitivity were investigated for the Antarctic marine diatom Chaetoceros brevis. Iron-limited and iron-replete cultures were exposed to identical daily irradiance levels, Supplied as dynamic (20-1350 mu mol m(-2) s(-1)) and constant (260 [mu mol m(-2) s(-1)) irradiance. After acclimation, growth, maximal quantum yield of PSII (F(v)/F(m)), pigment composition, and the activities of the antioxidant enzymes superoxide dismutase (SOD), ascorbate peroxidase (APX) and glutathione reductase (GR) were determined. Then, excessive irradiance sensitivity was assessed by monitoring pigment composition, F(v)/F(m) and viability loss during a single excessive PAR and UV treatment.Iron limitation reduced growth rates, F(v)/F(m), dynamics, and cellular pigments pools. Cellular pigiment concentrations were higher under dynamic irradiance than under constant irradiance but this difference was less pronounced under iron limitation compared to iron-replete conditions. SOD and APX activities increased during dynamic irradiance under iron limitation, suggesting increased radical formation around PSII Despite these physiological differences, no effects on growth were observed between constant and dynamic irradiance cultivation in iron-limited and iron-replete cells. The applied Culturing conditions did not affect glutathione reductase activity in C brevis. F(v)/F(m) and xanthophyll de-epoxidation dynamics during excessive irradiance were not different for iron-limited and replete cells and viability loss was not found during excessive irradiance. This Study revealed photoacclimation differences between iron-limited and iron-replete C brevis Cultures that did not affect growth rates and excessive irradiance sensitivity after acclimation to constant and dynamic irradiance. (C) 2008 Elsevier B.V. All rights reserved.

Antioxidant enzymes such as superoxide dismutase (SOD) play a key role in the removal of reactive oxygen species produced during visible and ultraviolet irradiance stress in microalgae and plants. However, little is known about the enzymatic antioxidative stress responses in ecologically important Antarctic marine microalgae. SOD in particular is difficult to analyze, possibly due to problems in obtaining sufficient quantities necessary for reliable and reproducible enzymatic assays. The aim of the present work was to create a sensitive, easy-to-use and reliable method for SOD determination in Antarctic microalgal material by comparing and optimizing existing protein extraction procedures and SOD assays in the marine Antarctic diatom Chaetoceros brevis. Optimization was achieved in cell disruption (sonication) and protein extraction procedures, extraction buffers, SOD assay methods (xanthine/xanthine oxidase and NBT/ riboflavin photometric quantitative methods and native gel electrophoresis qualitative method) and the assay temperature. Protein extraction was optimal at low sonication amplitudes after a few pulses, irrespective of the type of buffer used. Extraction efficiency varied highly between the tested buffers; most protein was extracted in the presence of 1% of Triton X-100. SOD activity was best quantified using the NBT/riboflavin method in combination with a buffer containing potassium phosphate and Triton X-100. Moreover, the NBT/ riboflavin method was demonstrated to be the most reliable and sensitive method at low temperatures (5 degrees C). (c) 2007 Elsevier B.V. All rights reserved.

The influence of photoacclimation on the effects of excessive photosynthetically active (PAR; 400-700 nm) and ultraviolet (UVR; 280-400 nm) radiation was assessed for the marine diatoms Thalassiosira weissflogii (Grunow) Fryxell and Hasle and Thalassiosira antarctica (Comber). Low and high PAR acclimated cultures were subjected to simulated surface irradiance (SSI) that mimicked irradiance around noon, including UVR. PSII efficiency, xanthophyll conversion, superoxide dismutase (SOD) activity, carbohydrate buildup, and lipid peroxidation were investigated after 30 min SSI and during 120 min recovery in low irradiance. Furthermore, viability loss was measured during 4 h SSI. Prior to SSI, the diadino-diatoxanthin pool was increased in high irradiance acclimated cells, compared with cells grown under low irradiance. Thirty-minutes SSI caused a pronounced decline in PSII efficiency. This coincided with de-epoxidation of diadinoxanthin in high irradiance acclimated cells, which was completely reversed during recovery in low irradiance. De-epoxidation was lower for low irradiance acclimated cells, whereas PSII efficiency and carbohydrate buildup were lower during the recovery phase. Furthermore, clear UVR effects on PSII efficiency were observed in low irradiance but not in high irradiance acclimated cells. Although 30 min SSI did not increase cellular SOD activity and lipid peroxidation, prolonged (4 h) SSI caused viability loss in low irradiance acclimated cells, which was enhanced by UVR. Therefore, PAR and UVR-induced PSII inactivation and viability loss were reduced by high irradiance-mediated changes in light harvesting and the xanthophyll pigments. In addition to photoacclimation-modulated differences, minor sensitivity differences were found between species.

Short-term photosensitivity and oxidative stress responses were compared for three groups of marine microalgae: Antarctic microalgae, temperate diatoms and temperate flagellates. In total, 15 low-light-acclimated species were exposed to simulated surface irradiance including ultraviolet radiation (SSI). Photosensitivity was assessed as the rate of recovery of Fv/Fm in the hours following SSI treatment. Before, during and after the SSI treatment, oxidative stress responses were assessed by following xanthophyll content and cycling, and activities of superoxide dismutase, ascorbate peroxidase and glutathione reductase, and glutathione redox status. When acclimated to low irradiance, antioxidant levels were not group specific. Superoxide dismutase activity was positively correlated with cell size, whereas in general, ascorbate peroxidase activity appeared to be lower and glutathione redox status appeared to be higher in the Antarctic than in the temperate species. After SSI exposure, the strong inhibition of PSII was followed by variable rates of recovery, although four species remained photosynthetically inactive. SSI tolerance appeared unrelated to geographic or taxonomic background, or to cell size. PSII recovery was enhanced in species with decreasing superoxide dismutase activity, glutathione redox status and increased xanthophyll cycle activity. We conclude that antioxidant responses are highly species specific and not related to the geographic or taxonomic background. Furthermore, xanthophyll cycling seems more important than antioxidants. Finally, it can be hypothesized that glutathione could function as a stress sensor and response regulator.

The wavelength dependency of xanthophyll cycling in two marine microalgae (Thalassiosira weissflogii and Dunaliella tertiolecta) was studied by establishing biological weighting functions (BWFs) during exposure to natural ultraviolet radiation. High-(HL) and low-(LL) light-acclimated cultures of both species were exposed outdoors for up to 60min under a series of UVR (280-400 nm) cut-off filters, after which the de-epoxidation state of xanthophyll cycle pigments, radiocarbon assimilation and photochemical quantum yield were measured. Exposures were repeated 4-8 times during the daily cycle to create exposure-response curves for each wavelength condition. UVR affected the three target processes significantly in both species and biological weights increased with decreasing wavelength, particularly in the UVBR region (280-315 nm). Minor wavelength dependency was observed between 315 and 360 nm. After BWF normalization to 300 nm, the LL cultures showed highly similar responses when comparing the three target processes, while the BWFs for the HL cultures differed significantly. The observed enhanced xanthophyll cycling activity in the UVR region implied that xanthophylls had an active role in diminishing UVR stress. However, this enhancement seems to be an indirect effect of damage within the dark reactions of photosynthesis. Hence, another vital target process further downstream in the photosynthetic process, possibly involved in the dark reactions, seems to be responsible for the high similarity in BWFs. Fil: Buma, Anita G. J.. University of Groningen; Países Bajos Fil: Visser, Ronald J. W.. University of Groningen; Países Bajos Fil: van de Poll, Willem H.. University of Groningen; Países Bajos Fil: Villafañe, Virginia Estela. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Fundación Playa Unión. Estación de Fotobiología Playa Unión; Argentina Fil: Janknegt, Paul J.. University of Groningen; Países Bajos Fil: Helbling, Eduardo Walter. Fundación Playa Unión. Estación de Fotobiología Playa Unión; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

Photoacclimation properties were investigated in two marine microalgae exposed to four ambient irradiance conditions: static photosynthetically active radiation (PAR: 400–700 nm), static PAR + UVR (280–700 nm), dynamic PAR and dynamic PAR + UVR. High light acclimated cultures of Thalassiosira weissflogii and Dunaliella tertiolecta were exposed outdoors for a maximum of 7 days. Dynamic irradiance was established by computer controlled vertical movement of 2 L bottles in a water filled basin. Immediate (

During austral summer 2006, experiments were carried out to evaluate the effects of ultraviolet radiation (UVR, 280-400 nm) on carbon fixation of natural phytoplankton assemblages from Patagonia (Argentina). Surface water samples were collected (ca. 100 m offshore) at mid morning using an acid-cleaned (1 N HCl) dark container. The short-term impact of UVR (measured as radiocarbon incorporation) was immediately assessed by exposing samples to three artificial illumination treatments: PAR (400-700 nm), PAR + UV-A (320-700 nm), and PAR + UV-A + UV-B (280-700 nm). Pico-nanoplankton characterized the assemblages, and taxon-specific pigment fingerprinting combined with CHEMTAX and supplemented with microscopic observations showed varied proportions of diatoms, chlorophytes, and cyanobacteria throughout January-February 2006. Photosynthetic efficiency, as assessed through assimilation numbers, was high [between 4.4 and 10.4 μg C (μg chl-a)-1 h-1], and it was probably favored by the supply of inorganic nutrients from the Chubut River. UVR-induced photoinhibition appeared to be related to the taxonomic composition: in general, higher photoinhibition was observed when diatoms dominated, whereas this was lower when samples were dominated by chlorophytes. Our data suggest that xanthophyll pigments might have provided only limited protection in these already highlighted acclimated assemblages. © 2008 Springer-Verlag. Fil: Villafañe, Virginia Estela. Fundación Playa Unión; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Janknegt, Paul J.. University of Groningen; Países Bajos Fil: de Graaff, Marco. University of Groningen; Países Bajos Fil: Visser, Ronald J. W.. University of Groningen; Países Bajos Fil: van de Poll, Willem H.. University of Groningen; Países Bajos Fil: Buma, Anita G. J.. University of Groningen; Países Bajos Fil: Helbling, Eduardo Walter. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Fundación Playa Unión; Argentina