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The red macroalgae Hydropuntia cornea, Gracilariopsis longissima and Halopithys incurva were cultured for 14 d under laboratory conditions, in enriched seawater with a high nutrient content (N-NH4+ and P-PO43- ) and two radiation regimes: PAR (400-700 nm) and PAB (280-700 nm). The UV radiation effects under high availability of nutrients on growth, photosynthetic pigments (chlorophyll a, carotenoids and phycobiliproteins), photosynthetic activity and biochemical composition were studied. Maximum quantum yield (Fv /Fm ) was not significantly different among the PAR and PAB treatments during the experiment. However, the maximum electronic transport rate (ETRmax ) increased over time, showing the highest values in PAR for H. incurva and H. cornea, whereas for G. longissima it was found in PAB. Photosynthetic efficiency (αETR ) decreased over time in the first two species, but increased in G. longissima. Saturation irradiance (EkETR ) and maximum nonphotochemical quenching (NPQmax ) increased in PAB with time up to 80% and 30%, respectively, indicating a photosynthetic acclimatization like that of sun-type algae. Five MAAs were identified in all species using high performance liquid chromatography (HPLC). The total content of MAAs increased over time, being 30% higher in H. incurva, 40% in G. longissima and 50% in H. cornea in PAB than in the PAR treatment. Finally, the antioxidant activity was also higher in the PAB treatment. All of the species presented an effective mechanism of photoprotection based on the accumulation of photoprotective compounds with antioxidant activity, as well as a high dissipation of excitation energy (high NPQmax ).

The culture of microalgae and cyanobacteria in open systems has been improved through the novel approach of thin-layer raceway ponds. The importance of studying mass cultivation of the cyanobacterium Nostoc calcicola (further as Nostoc) lies in its biotechnological potential as a source of bioactive compounds for food and non-food applications. These compounds include polysaccharides, mycosporine-like amino acids and phycocyanin. Nostoc was cultured outdoors in a thin-layer raceway pond where the biomass production, physiological status, photosynthetic activity, and biochemical composition were monitored through the experimental period of 5 days. The biomass, as did the maximal quantum yield of PSII, maximal electron transport rate (ETRmax) and photosynthetic efficiency (αETR) increased throughout the experimental period showing the optimal operation of the thin-layer raceway ponds, due to the light penetrates deeper into the thin culture layer and thus more light is available to the cells. Oxygen levels in the culture increased over time, but no photoinhibition was evident indicating optimal action of non-photochemical mechanisms. Nostoc increased the total internal carbon content over the experimental period. Chlorophyll increased, whereas the N compounds such as the biliprotein phycocyanin decreased. Among the UV-absorbing compounds, polyphenols, mycosporine-like amino acids, such as shinorine and other unknown UV-A absorbing compounds were detected. There components showed a positive correlation to antioxidant activity. Thus, the optimal accumulation of biomass and the accumulation of bio-active compounds having antioxidant capacity show the possible biotechnological applications of Nostoc.

Electrical lighting favours the development of photosynthetic biofilms in caves which can induce biodeterioration in the colonized substrates. The use of specific lights as a limiting factor for biofilm growth could be effective in their control and represents an alternative to chemical methods since they can damage the substrate. However, studies about lighting and the photosynthetic activity of organisms in caves are scarce. In order to select the most effective LED light source in reducing photosynthesis and therefore, in reducing the growth rates of microalgae and cyanobacteria, four biofilms in the Nerja Cave were illuminated by several light emitted diodes (LEDs) with different spectral compositions and the photobiological responses were measured both by empirical and theoretical methodologies. The empirical approach was based on the photosynthetic efficiency, by measuring the in vivo chlorophyll a (Chl a) fluorescence and the theoretical approach was based on the photonic assimilation performance related to the proportion of the light quality used for photosynthesis, according to the action spectra for photosynthesis available in the literature. The photobiological responses showed differences between the empirical and theoretical approach mainly in biofilms dominated by cyanobacteria and red algae, probably because the available action spectra were not useful for monitoring these Nerja Cave biofilms. However, the expected spectral responses of photosynthesis were observed in green microalgal biofilms with maximum photosynthetic efficiency in red and blue light although the green light was also unexpectedly high. The high photosynthetic efficiency in green light could be explained by the predictable high chlorophyll content due to a very dark environment. The results were not conclusive enough for all the biofilm types to be able to recommend a specific lighting system for the photocontrol of biofilm expansion. Therefore, new action spectra for photosynthesis of the extremophile organisms of the Nerja Cave are required. This approach, based on theoretical and empirical methodologies, is a useful tool to obtain information to allow the design of the most adequate lighting systems to reduce photosynthetic activity and favour the conservation of the caves.

The combined effects of ultraviolet radiation (UVR) and nutrients on the red macroalga Gracilariopsis longissima were analyzed for 14 days in a 2 × 2 factorial experiment design. Cell ultrastructure, photosynthetic activity as in vivo chlorophyll a fluorescence and the accumulation of bioactive compounds as soluble proteins, starch, lipids, polyphenols and mycosporine-like amino acids (MAAs) were evaluated. Three main morphological alterations as defense mechanisms against UVR were observed, i.e., an increase in cell wall thickness, accumulation of floridean starch grains and multiplication of the number of layers of cortical cells. Interaction of light and nutrients affected photosynthetic variables, such as electron transport rate (estimator of photosynthetic capacity) and photoinhibition irradiance (E opt ), and also the level of N compounds (i.e., proteins and MAAs) and total carbon and polyphenol content. Other variables such as chlorophyll a , carotenoids, starch, lipids, antioxidant activity and cell viability were influenced by both factors, or only by light, but without interactive effects. G . longissima showed a high acclimation to elevated UVR and nutrients by the decrease of the photosynthetic rate and the increase in the accumulation of N compounds with photoprotection and antioxidant capacities. On the other hand, starch, lipid and polyphenols increased mainly in the presence of UVR under low nutrient availability. The biotechnological implications of the metabolic changes due to N availability and UVR in this species are discussed.

In order to evaluate the photosynthetic efficiency of cave photoautotrophic biofilms, in vivo chlorophyll a fluorescence was measured using two different fluorometers, in two biofilms in the touristic karstic Nerja Cave (Spain). The study was done for entire days in summer and in winter during the first year and repeated for a second year, in order to cover the widest range of environmental conditions, i.e., atmospheric CO2, temperature, seeping water and relative humidity levels. Effective quantum yield and relative electron transport rate (rETR) were determined during periods of light whereas maximal quantum yield (Fv/Fm) was determined in situ during dark periods. On summer days, in situ photosynthetic yields in cyanobacterium biofilms (Chroococcidiopsis sp.) increased 7–16 times compared to that of winter days, whereas in biofilms comprised of green and red microalgae and various cyanobacterium species, no seasonal or yearly variations were observed. In contrast, maximal rETR in the two biofilms increased in the periods with the highest values of both CO2 and relative humidity. Positive correlations between all environmental variables and rETR were found. According to Redundancy Analysis, all environmental variables, mainly CO2 and relative humidity were related to photosynthetic variables. The effective quantum yields showed different values depending on the measuring light of the PAM. The values were higher with red light (Diving PAM) compared to blue light (Junior PAM) mainly in the site dominated by cyanobacteria. Nerja Cave is shown as an excellent place to study the effects of light and CO2, among other environmental variables of biofilm photosynthetic activity. The monitoring of photosynthetic activity by in vivo chlorophyll a fluorescence could be used to follow the effects of the treatments applied by the touristic cave managers to reduce the proliferation of biofilms composed of various species, and, consequently, the biodeterioration of speleothems could be reduced.

This study was designed to evaluate the potential use of algal extracts in cosmeceuticals, including factors related to biosecurity. The aqueous crude extracts of Hydropuntia cornea and Gracilariopsis longissima showed a good photoprotective capacity (Sun Protection Factor, SPF) due to, among other reasons, the presence of five types of mycosporine-like amino acids (MAAs) detected by high pressure liquid chromatography-photodiode array detector (HPLC-PDA) and electrospray ionization mass spectrometry (ESI-MS) (Palythine, Asterina-330, Shinorine, Porphyra-334, and Palythinol). The toxicity of the extracts was evaluated by the MTT assay, which is based on the metabolic reduction of MTT [3-(4,5-dimethylthiazol-2yl)-diphenyl tetrazolium bromide] by the action of the mitochondrial enzyme succinate dehydrogenase. This assay was carried out in vitro in three cell lines: one related to the immune system (murine macrophages of the immune system: RAW264.7) and two human cell lines related to the skin (gingival fibroblasts: HGF, and immortalized human keratinocytes: HaCaT). Both extracts showed no cytotoxic activity in both types of human cells, whereas they showed cytotoxicity in murine tumor cells of the immune system (macrophages: RAW264.7). On the other hand, the immunological activity in the murine macrophage RAW264.7 was studied at a concentration lower than 100 &mu, g mL&minus, 1 and lower than the EC50, and evaluated by the production of pro-inflammatory compounds through an immunosorbent assay linked to enzymes such as tumor necrosis factor-&alpha, (TNF-&alpha, ) or anti-inflammatory/proinflammatory enzymes such as interleukin-6 (IL-6). Both algae extracts induced the biosynthesis of TNF-&alpha, and IL-6. The production of TNF-&alpha, was much higher than that observed in the control (at a concentration of the aqueous extract higher than 5 &mu, 1). These results support the theory that the extracts of H. cornea and G. longissima actively induce the production of cytokines. In summary, the extracts of these species did not show cytotoxicity in human cells, and they present with immunomodulatory and photoprotection capacity.

Acanthophora spicifera (M.Vahl) Borgesen is a macroalga of great economic importance. This study evaluated the antioxidant responses of two algal populations of A. spicifera adapted to different abiotic conditions when exposed to ultraviolet-A+ultraviolet-B radiation (UV-A+UV-B). Experiments were performed using the water at two collection points for 7 days of acclimatization and 7 days of exposure to UVR (3 h per day), followed by metabolic analyses. At point 1, water of 30 ± 1 practical salinity unit (psu) had concentrations of 1.06 ± 0.27 mm NH 4 + , 8.47 ± 0.01 mm NO 3 - , 0.17 ± 0.01 mm PO 4 - 3 and pH 7.88. At point 2, water of 35 ± 1 psu had concentrations of 1.13 ± 0.05 mm NH 4 + , 3.73 ± 0.01 mm NO 3 - , 0.52 ± 0.01 mm PO 4 - 3 and pH 8.55. Chlorophyll a, phycobiliproteins, carotenoids, mycosporins, polyphenolics and antioxidant enzymes (catalase, superoxide dismutase and guaiacol peroxidase) were evaluated. The present study demonstrates that ultraviolet radiation triggers antioxidant activity in the A. spicifera. However, such activation resulted in greater responses in samples of the point 1, with lower salinity and highest concentration of nutrients.

The safety and stability of synthetic UV-filters and the procedures for evaluating the photoprotective capability of commercial sunscreens are under continuous review. The influence of pH and temperature stressors on the stability of certain Mycosporine-like amino acids (MAAs) isolated at high purity levels was examined. MAAs were highly stable at room temperature during 24 h at pH 4.5&ndash, 8.5. At 50 &deg, C, MAAs showed instability at pH 10.5 while at 85 &deg, C, progressive disappearances were observed for MAAs through the studied pH range. In alkaline conditions, their degradation was much faster. Mycosporine-serinol and porphyra-334 (+shinorine) were the most stable MAAs under the conditions tested. They were included in four cosmetically stable topical sunscreens, of which the Sun Protection Factor (SPF) and other Biological Effective Protection Factors (BEPFs) were calculated. The formulation containing these MAAs showed similar SPF and UVB-BEPFs values as those of the reference sunscreen, composed of synthetic UV absorbing filters in similar percentages, while UVA-BEPFs values were slightly lower. Current in vitro data strongly suggest that MAAs, as natural and safe UV-absorbing and antioxidant compounds, have high potential for protection against the diverse harmful effects of solar UV radiation. In addition, novel complementary in vitro tests for evaluation of commercial sunscreens efficacy are proposed.

Dissolved oxygen concentration is a critical point for microalgae in large scale cultivation systems. Highly productive cultures inevitably generate a build-up of oxygen gradients along the reactor which can affect photosynthetic performance. In this study, a fast-growing strain of Chlorella vulgaris R117 was cultured outdoors in a thin-layer cascade during a one-week trial reaching a biomass productivity of 4 g DW L−1 d−1 (27 g DW m−2 d−1). High photosynthetic activity led to oxygen oversaturation of up to 400% in some parts along the culture units at midday. The aim was to examine the effect of high dissolved oxygen concentration on diurnal changes in the photosynthetic performance and growth of the Chlorella culture using multi-technique approach. Photosynthetic activity of Chlorella R117 culture was estimated in situ and ex situ using oxygen production and in vivo Chl a fluorescence measurements, which showed good correlation. The rates of electron transport and of oxygen production were related, but the values of the μmolelectrons/μmolO2 ratio was higher than predicted, suggesting the probable involvement of electron and oxygen consuming processes such as photorespiration and Mehler reaction. These processes probably function as photoprotective mechanisms, since no photodamage was observed in the Chlorella R-117 cultures. Depression (down-regulation) of photosynthetic activity due to the exposition to high dissolved oxygen concentration along the cascade area over time was observed. The usefulness of on-line measurements was demonstrated to obtain immediate and in-situ information on the physiological status of the culture. This data can be used in models of operation control for large-scale microalgae production units.

Photosynthetic activity was estimated, in 2 different sets of experiments, as in vivo chlorophyll a fluorescence in 2 calcareous red macroalgae: Neogoniolithon brassica-florida, which forms vermetid reefs with Dendropoma petraeum, and the rhodolith-forming Lithophyllum margaritae. In the first set of experiments, vermetid reefs collected from the Cabo de Gata-Nijar Natural Park (Almeria, Spain) were incubated for 6 months in outdoor mesocosms at 2 different temperatures: ambient temperature (T) and ambient temperature plus 2 oC (T+). Daily variations in effective quantum yields were related to the increase in solar radiation. Electron transport rate (ETR) was higher at T+ than at T. Oxygen production, estimated from ETR, was higher in N. brassica-florida growing in vermetid reefs in the coastal area than in those incubated in mesocosms or under laboratory conditions. In the second set of experiments, L. margaritae specimens with or without attached epifauna (mainly sponges) were collected at 2 sites (Pedregoso and CFE) on the Baja California Peninsula, Mexico, and at 2 depths (1 and 2 m). ETR values for rhodoliths collected at 1 m depth and with attached epifauna were higher than those for rhodoliths collected at 2 m depth, indicating the importance of greater incident radiation on photosynthetic activity. Photosynthetic activity was higher in L. margaritae collected from Pedregoso, the site with better water quality (high transparency and low pollution) and rocky substrate availability. In the CFE samples, the photosynthetic efficiency (alphaETR) was highest in rhodoliths with attached epifauna. The highest ETR values for N. brassica-florida at T+ and the highest alphaETR or ETR values for L. margaritae with attached sponges may be explained by the increase in CO2 available for photosynthesis due to the respiration of the associated fauna. The usefulness of in vivo chlorophyll a fluorescence for estimating photosynthetic production in calcareous macroalgae under multifactorial experiments is discussed. La actividad fotosintetica fue estimada, en 2 series experimentales, mediante la fluorescencia in vivo de la clorofila a en 2 algas rojas calcareas: Neogoniolithon brassica-florida, que forma arrecifes de vermetidos con Dendropoma petraeum, y Lithophyllum margaritae (rodolitos). En la primera serie, vermetidos recolectados en el Parque Natural Cabo de Gata-Nijar (Almeria, Espana) fueron incubados en mesocosmos a diferentes temperaturas: temperatura ambiental (T) y temperatura ambiental mas 2 oC (T+). Las variaciones diurnas en el rendimiento cuantico efectivo estuvieron relacionadas con el incremento de irradiancia solar. La tasa de transporte de electrones (ETR) fue mas alta en el tratamiento T+ que en T. La produccion de oxigeno, estimada a partir de la ETR, fue mayor en N. brassica-florida del medio natural que en vermetidos incubados en mesocosmos o en el laboratorio. En la segunda serie experimental, L. margaritae con o sin epifauna (dominada por esponjas) fue recolectada en 2 sitios (Pedregoso y CFE) de la peninsula de Baja California, Mexico, a 2 profundidades (1 y 2 m). La ETR de rodolitos recolectados a 1 m de profundidad con epifauna asociada fue mas alta que la de aquellos recolectados a 2 m de profundidad, indicando la importancia del incremento de la irradiancia sobre la produccion fotosintetica. La actividad fotosintetica fue mayor en L. margaritae de Pedregoso, el lugar con mejor calidad de agua (alta trasparencia y baja contaminacion) y con disponibilidad de sustrato rocoso. En CFE, la eficiencia fotosintetica (alfaETR) fue mas alta en los rodolitos con epifauna. Los valores mas altos de ETR para N. brassica-florida a T+ y de alfaETR o ETR para L. margaritae con esponjas fijadas podrian ser explicados por el incremento de CO2 disponible para la fotosintesis procedente de la respiracion de la fauna asociada. En este trabajo se discute la utilidad de la fluorescencia in vivo de la clorofila a para estimar la fotosintesis en macroalgas calcareas en experimentos multifactoriales.

Methanolic and hydroethanolic extracts of ten common species of marine macroalgae (5 Rhodophyta, 2 Ochrophyta and 3 Chlorophyta) of the intertidal system of the Canary Islands were screened to evaluate their in vitro antioxidant activity by DPPH radical scavenging activity and reducing power. The concentration of bioactives as pigments and phenolic compounds was also determined. Wild-collected algae and these cultivated in an integrated multitrophic aquaculture (IMTA) system during one month, were analyzed. The highest antioxidant activity was observed in Halopithys incurva, Fucus spiralis and Treptacantha abies-marina. The hydroethanolic extracts exhibited higher reducing power than methanolic extracts in most species. However, the differences in DPPH scavenging activity between solvents were not so high, and only the hydroethanolic extracts of some species showed antioxidant activity. The highest concentration of polyphenols was reached in H. incurva, T. abies-marina and F. spiralis. However, compared to collected seaweeds, the percentage of these compounds was lower in cultivated samples. The highest concentrations of chlorophyll a and carotenoids were found in T. abies-marina, Codium intertextum and F. spiralis. In red algae, the phycoerythrin (PE) content was higher in H. incurva, followed by P. capillacea. After culture, pigment content increased in most species, depending on the assimilation capacity of the ammonium from fishpond effluents. C/N ratio in samples collected in the field was high, indicating nitrogen limitation. After cultivation under high nitrogen availability (fishpond effluents), the ratio decreased in all species, especially in C. intertextum, F. spiralis and H. incurva, by the accumaltion of N-compounds. Pearson coefficient only showed significant correlations between the reducing power and some bioactive compounds, particularly polyphenols and PE. In conclusion, culture under IMTA conditions changed the biochemical composition of algae. The antioxidant activity did not decrease after culture in several species, despite the reduction of phenolic compounds, probably due to the benefits of the high N availability in the IMTA.

The red macroalgae Hydropuntia cornea, Gracilariopsis longissima and Halopithys incurva were cultured for 14 d under laboratory conditions, in enriched seawater with a high nutrient content (N-NH

This chapter presents the use of both macro- and microalgae for bioremediation of different effluents such as from fishponds, urban sewage treatments, and pig farms. The biofiltration of nitrogen and phosphate compounds by algae is followed by the determination of nutrient uptake rate and nutrient uptake efficiency. In the case of macroalgae, biofiltration of fishpond effluents are discussed in the frame of integrated multitrophic aquaculture approaches. In the case of urban sewage and pig slurry, microalgae growing in race-ways or in thin layer cascade cultivators are used to reduce nutrient levels, so that the effluents have sufficient quality to be discharged to the environment. The use of macro- and microalga biomass grown in the effluents as a source of biocompounds with cosmetic and nutraceutic interest is discussed. Different bioassays are included with antioxidant and immunostimulant capacities. The economic and ecological benefits of the bioremediation and biorefinery by using algae are discussed.

This study evaluates the production of biomass and mycosporine-like amino acids (MAAs) throughout the year in Gracilaria vermiculophylla (Rhodophyta) collected in Ria de Aveiro (Portugal). The algae were grown in outdoor tanks in seawater with the addition of fishpond effluents under two different water flows (100 and 200 L h−1) in an integrated multi-trophic aquaculture (IMTA) system (tanks 1200 L; 1.5 m2) and different algal densities (3, 5, and 7 kg m−2). MAA content in IMTA seaweeds was significantly affected by the interaction of time and stocking density, but not by the water flow. The highest MAA content was observed in April (about 3.13 mg g−1 DW) followed by May (1.79 mg g−1 DW). Seaweed biomass productivity was higher in May (372.06 g DW m−2 week−1) than in April (353.40 g DW m−2 week−1). Four MAAs were identified by HPLC and electrospray ionization mass spectrometry (ESI-MS) in G. vermiculophylla: Porphyra-334, Shinorine, Palythine and Asterina-330. The highest levels of Porphyra-334 and Shinorine were reached from November to January and the Palythine + Asterina-330 from April to August. Taking into account the average biomass and MAA production of G. vermiculophylla growing in this IMTA system (8.56 g of MAA in 18 m2 culture along 8 months; 35.5% produced in April), a total amount of 71.33 g MAA year−1 could be produced in this system by scaling up to 100 m2. MAAs could be further used as photoprotector and antioxidant compounds in cosmetic products.

Up to now, scarce information has been available regarding major photobiochemical changes that accompany the metabolic shift from heterotrophy to phototrophy in microalgae cells. In the present trials the trophic conversion from heterotrophic to phototrophic growth regime was studied in the microalga Chlorella vulgaris g120 grown in outdoor thin-layer cascades. The crucial question was whether Chlorella g120 can undergo the photoacclimation of its photosynthetic apparatus under natural irradiance. As for comparison the phototrophic strain Chlorella vulgaris R-117 was cultured in parallel. Various in-situ and ex-situ monitoring techniques namely oxygen production and chlorophyll fluorescence were tested to characterize physiological response of the Chlorella g120 culture during photoacclimation and correlate it with growth. We show that Chlorella g120 can undergo the metabolic shift from heterotrophic to phototrophic growth regime, but its conversion is. A rather high respiration rate was interpreted as a strategy to dissipate the unused light energy. Chlorella g120 was found as the strain characterized by a small-antenna size strain (low chlorophyll/cell ratio The build-up of dissolved oxygen (DO) concentration and high electron transport rate measured in-situ indicated high photosynthetic activity of outdoor microalgae cultures. These variables are usable on-line as reliable markers as they reflect the physiological status of microalgae culture. Highly productive cultures, like Chlorella R-117 have high photosynthesis which is inevitably accompanied by the build-up on high DO gradients. To maintain high biomass production in such strains we have to find an interplay among photosynthetic activity, gradients of DO concentration, biomass density and culture layer thickness that can secure high growth rate (and productivity).