A Blend Consisting of Agaran from Seaweed Gracilaria birdiae and Chromium Picolinate Is a Better Antioxidant Agent than These Two Compounds Alone.

A blend refers to the combination of two or more components to achieve properties that are superior to those found in the individual products used for their production. Gracilaria birdiae agaran (SPGb) and chromium picolinate (ChrPic) are both antioxidant agents. However, there is no documentation of blends that incorporate agarans and ChrPic. Hence, the objective of this study was to generate blends containing SPGb and ChrPic that exhibit enhanced antioxidant activity compared to SPGb or ChrPic alone. ChrPic was commercially acquired, while SPGb was extracted from the seaweed. Five blends (B1; B2; B3; B4; B5) were produced, and tests indicated B5 as the best antioxidant blend. B5 was not cytotoxic or genotoxic. H2O2 (0.6 mM) induced toxicity in fibroblasts (3T3), and this effect was abolished by B5 (0.05 mg·mL-1); neither ChrPic nor SPGb showed this effect. The cells also showed no signs of toxicity when exposed to H2O2 after being incubated with B5 and ChrPic for 24 h. In another experiment, cells were incubated with H2O2 and later exposed to SPGb, ChrPic, or B5. Again, SPGb was not effective, while cells exposed to ChrPic and B5 reduced MTT by 100%. The data demonstrated that B5 has activity superior to SPGb and ChrPic and points to B5 as a product to be used in future in vivo tests to confirm its antioxidant action. It may also be indicated as a possible nutraceutical agent.

Production of galactan phthalates derivatives extracted from Gracilaria birdie: Characterization, cytotoxic and antioxidant profile.

The present work explores the esterification reaction in the polysaccharide extracted from the seaweed Gracilaria birdiae and investigates its antioxidant potential. The reaction process was conducted with phthalic anhydride at different reaction times (10, 20 and 30 min), using a molar ratio of 1:2 (polymer: phthalic anhydride). Derivatives were characterized by FTIR, TGA, DSC and XRD. The biological properties of derivatives were investigated by assays of cytotoxicity and antioxidant activity (2,2-diphenyl-1-picrylhydroxyl - DPPH and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt - ABTS). The results obtained by FT-IR confirmed the chemical modification, there was a reduction related to the presence of carbonyl and hydroxyl groups when compared to the in nature polysaccharide spectrum. TGA analysis showed a change in the thermal behavior of the modified materials. X-ray diffraction, it was shown that the in nature polysaccharide appeared as an amorphous material, while the material obtained after the chemical modification process had increased crystallinity, due to the introduction of phthalate groups. For the biological assays, it was observed that the phthalate derivative was more selective than the unmodified material for the murine metastatic melanoma tumor cell line (B16F10), revealing a good antioxidant profile for DPPH and ABTS radicals.

Heavy Metal Depuration Steps for Gracilaria chilensis in Outdoor Culture Systems.

Seaweed aquaculture is affected by natural and anthropogenic stressors, which put the biomass productivity of the cultures at risk. Seaweed biomass for commercial purposes, principally in pharmaceutical and/or nutraceutical applications, needs to be free of pollutants; therefore, controlled cultures have relevance in regulating the quality of biomass. The aim of this work was to demonstrate the successful utilization of controlled outdoor cultures to remove excess heavy metal accumulation in Gracilaria chilensis, an important commercial seaweed farming model. Specifically, we designed a simple and operational heavy metal depuration protocol, utilizing seawater and tap water removal, which permitted the concentration reduction of 10 heavy metals, including As, Cu, and Cd but not Zn, from the biomass at 7 days of culture. The percentage of depuration of the heavy metals ranged from 32 to 92% at 7 days, which was maintained throughout 21 days of culture. During the culture period, the monitored physicochemical parameters (temperature, salinity, and dissolved oxygen, among others) remained stable, with an increase in the daily growth rate (DGR% d-1) of the biomass recorded after 14 days of culture. Consequently, the experimental setup was successful for heavy metal depuration, which highlights the importance of controlled outdoor cultures as important tools of sustainability.

Pre-domestication bottlenecks of the cultivated seaweed Gracilaria chilensis.

Gracilaria chilensis is the main cultivated seaweed in Chile. The low genetic diversity observed in the Chilean populations has been associated with the over-exploitation of natural beds and/or the founder effect that occurred during post-glacial colonization from New Zealand. How these processes have affected its evolutionary trajectory before farming and incipient domestication is poorly understood. In this study, we used 2232 single nucleotide polymorphisms (SNPs) to assess how the species' evolutionary history in New Zealand (its region of origin), the founder effect linked to transoceanic dispersion and colonization of South America, and the recent over-exploitation of natural populations have influenced the genetic architecture of G. chilensis in Chile. The contrasting patterns of genetic diversity and structure observed between the two main islands in New Zealand attest to the important effects of Quaternary glacial cycles on G. chilensis. Approximate Bayesian Computation (ABC) analyses indicated that Chatham Island and South America were colonized independently near the end of the Last Glacial Maximum and emphasized the importance of coastal and oceanic currents during that period. Furthermore, ABC analyses inferred the existence of a recent and strong genetic bottleneck in Chile, matching the period of over-exploitation of the natural beds during the 1970s, followed by rapid demographic expansion linked to active clonal propagation used in farming. Recurrent genetic bottlenecks strongly eroded the genetic diversity of G. chilensis prior to its cultivation, raising important challenges for the management of genetic resources in this incipiently domesticated species.

Insights into agar and secondary metabolite pathways from the genome of the red alga Gracilaria domingensis (Rhodophyta, Gracilariales).

Gracilariales is a clade of florideophycean red macroalgae known for being the main source of agar. We present a de novo genome assembly and annotation of Gracilaria domingensis, an agarophyte alga with flattened thallus widely distributed along Central and South American Atlantic intertidal zones. In addition to structural analysis, an organizational comparison was done with other Rhodophyta genomes. The nuclear genome has 78 Mbp, with 11,437 predicted coding genes, 4,075 of which did not have hits in sequence databases. We also predicted 1,567 noncoding RNAs, distributed in 14 classes. The plastid and mitochondrion genome structures were also obtained. Genes related to agar synthesis were identified. Genes for type II galactose sulfurylases could not be found. Genes related to ascorbate synthesis were found. These results suggest an intricate connection of cell wall polysaccharide synthesis and the redox systems through the use of L-galactose in Rhodophyta. The genome of G. domingensis should be valuable to phycological and aquacultural research, as it is the first tropical and Western Atlantic red macroalgal genome to be sequenced.

Characterization of an Agarophyton chilense Oleoresin Containing PPARγ Natural Ligands with Insulin-Sensitizing Effects in a C57Bl/6J Mouse Model of Diet-Induced Obesity and Antioxidant Activity in Caenorhabditis elegans.

The biomedical potential of the edible red seaweed Agarophyton chilense (formerly Gracilaria chilensis) has not been explored. Red seaweeds are enriched in polyunsaturated fatty acids and eicosanoids, which are known natural ligands of the PPARγ nuclear receptor. PPARγ is the molecular target of thiazolidinediones (TZDs), drugs used as insulin sensitizers to treat type 2 diabetes mellitus. Medical use of TZDs is limited due to undesired side effects, a problem that has triggered the search for selective PPARγ modulators (SPPARMs) without the TZD side effects. We produced Agarophyton chilense oleoresin (Gracilex®), which induces PPARγ activation without inducing adipocyte differentiation, similar to SPPARMs. In a diet-induced obesity model of male mice, we showed that treatment with Gracilex® improves insulin sensitivity by normalizing altered glucose and insulin parameters. Gracilex® is enriched in palmitic acid, arachidonic acid, oleic acid, and lipophilic antioxidants such as tocopherols and ß-carotene. Accordingly, Gracilex® possesses antioxidant activity in vitro and increased antioxidant capacity in vivo in Caenorhabditis elegans. These findings support the idea that Gracilex® represents a good source of natural PPARγ ligands and antioxidants with the potential to mitigate metabolic disorders. Thus, its nutraceutical value in humans warrants further investigation.

Salinity tolerance of macroalgae Gracilaria birdiae / Tolerância à salinidade da macroalga Gracilaria birdiae

This study aimed to determining the manner in which salinity influenced the growth of the macroalgae Gracilaria birdiae, with the objective of identifying its tolerance limits to this abiotic variable. The specimens were submitted to nutrient-enriched water of varying salinities (0, 10, 20, 30, 40, 50, and 60 ppt) for a 30-day period. Initially, under extreme salinity conditions (0 and 60 ppt) the growth of the G. birdiae suffered a negative impact. The macroalgae biomass exposed to 0 and 10 ppt salinities showed a reduction from day six until the experiment was completed. The macroalgae biomass exposed to salinities 20, 30, 40, and 50 ppt showed an increase, with no significant differences between the four treatments. This suggested that this salinity range was comfortable for this species to develop. We concluded that salinity is a crucial parameter which controls the growth of the G. birdiae. This seaweed was negatively influenced when exposed to values equal to or below 10 ppt and equal to 60 ppt, demonstrating good tolerance to salinities of 20, 30, 40 and 50 ppt.(AU)

Objetivamos com o presente trabalho avaliar os efeitos da salinidade sobre o crescimento da macroalga Gracilaria birdiae, visando identificar os seus limites de tolerância a esta variável abiótica. Os exemplares foram submetidos a água com salinidades 0, 10, 20, 30, 40, 50 e 60 ppt enriquecida com nutrientes por um período de 30 dias. Inicialmente a G. birdiae foi negativamente afetada em condições de salinidades extremas (0 e 60 ppt). A biomassa das macroalgas expostas a salinidade 0 e 10 ppt declinou a partir do 6º dia até o final do experimento. Houve aumento na biomassa das macroalgas expostas as salinidades 20, 30, 40 e 50 ppt, não apresentando diferenças significativas entre estes quatro tratamentos, sugerindo este intervalo de salinidade como sendo ótimo para o desenvolvimento desta espécie. Concluímos que a salinidade é um parâmetro importante para controlar o crescimento da G. birdiae, sendo afetada negativamente quando exposta a valores igual ou menor que 10 ppt e igual a 60 ppt, possuindo tolerância às salinidades de 20, 30, 40 e 50 ppt.(AU)

Sulfated polysaccharide from Gracilaria caudata reduces hypernociception and inflammatory response during arthritis in rodents.

Polysaccharide from marine alga Gracilaria caudata has potential health benefits, such as anti-inflammatory, gastroprotective and antidiarrheal effects. Here, we investigated the effect of a sulfated polysaccharide from G. caudata (SP-GC) on hypernociception and inflammatory response in arthritis models. The animals received SP-GC (3, 10 or 30 mg/kg) 1 h before tibio-tarsal injection of zymosan. Hypernociception, histopathology, edema, vascular permeability, myeloperoxidase (MPO) activity, cell influx, interleukin (IL)-1ß and nitric oxide (NO) levels were evaluated in acute phase. In another protocol, animals received SP-GC (30 mg/kg) 2 h post-complete Freund's adjuvant (CFA). Hypernociception, edema and arthritis index were determined in acute, sub-chronic and chronic phases. Rota-rod test measured the motor performance. SP-GC significantly reduced, in a dose-dependent manner, the zymosan-induced hypernociception with maximal effect at 30 mg/kg. The microscopic inflammation, joint edema, MPO activity, cell influx, IL-1ß and NO levels were also reduced by SP-GC. In the CFA-induced arthritis, SP-GC inhibits the hypernociception, edema and arthritic index in acute, sub-chronic and chronic phases. SP-GC did not alter the motor performance of animals. In conclusion, SP-GC exerts protective effect in models of arthritis due to the modulation of cell influx, IL-1ß and NO levels, culminating in the reduction of hypernociception and edema.

Chemical structure, anti-inflammatory and antinociceptive activities of a sulfated polysaccharide from Gracilaria intermedia algae.

The present work aimed at carrying out the isolation and biochemical characterization of a sulfated polysaccharide fraction (PLS) from the marine algae Gracilaria intermedia and investigating its anti-inflammatory and antinociceptive potential. PLS was obtained through enzymatic digestion with papain and analyzed by means of gel permeation chromatography and Nuclear Magnetic Resonance to 1H and 13C. In order to evaluate the potential of anti-inflammatory action of PLS, we performed paw edema induced by carrageenan, dextran, compound 48/80, histamine and serotonin. In addition, we also measured the concentration of myeloperoxidase, cytokines, the count of inflammatory cells and performed tests of the nociception. The PLS isolated was of high purity and free of contaminants such as proteins, and had molecular weight of 410 kDa. The same macromolecule was able to decrease the paw edema induced by all inflammatory agents (P < 0.05), myeloperoxidase (MPO) activity, neutrophil migration and IL-1ß levels. It also decreased acetic acid-induced writhing (P < 0.05) and formalin-induced paw licking time (P < 0.05), but no in hot plate test. In summary, the PLS decreased the inflammatory response by reducing neutrophil migration and modulating IL-1ß production and antinociceptive effects by a peripheral mechanism dependent on the down-modulation of the inflammatory mediators.

Antioxidant Sulfated Polysaccharide from Edible Red Seaweed Gracilaria birdiae is an Inhibitor of Calcium Oxalate Crystal Formation.

The genus Gracilaria synthesizes sulfated polysaccharides (SPs). Many of these SPs, including those synthesized by the edible seaweed Gracilaria birdiae, have not yet been adequately investigated for their use as potential pharmaceutical compounds. Previous studies have demonstrated the immunomodulatory effects of sulfated galactans from G. birdiae. In this study, a galactan (GB) was extracted from G. birdiae and evaluated by cell proliferation and antioxidant tests. GB showed no radical hydroxyl (OH) and superoxide (O2-) scavenging ability. However, GB was able to donate electrons in two further different assays and presented iron- and copper-chelating activity. Urolithiasis affects approximately 10% of the world's population and is strongly associated with calcium oxalate (CaOx) crystals. No efficient compound is currently available for the treatment of this disease. GB appeared to interact with and stabilize calcium oxalate dihydrate crystals, leading to the modification of their morphology, size, and surface charge. These crystals then acquired the same characteristics as those found in healthy individuals. In addition, GB showed no cytotoxic effect against human kidney cells (HEK-293). Taken together, our current findings highlight the potential application of GB as an antiurolithic agent.

Polysaccharide from Gracilaria caudata protects the human esophageal mucosal barrier: A differential topical effect and structural dependence.

This study aimed to evaluate the in vitro protective effect of topical treatment with a native sulfated polysaccharide of G. caudata (SP-Gc), hydrolyzed (H-SP-Gc), or desulfated (D-SP-Gc) polysaccharide of Gracilaria caudata in esophageal biopsies obtained from GERD patients. Biopsies were obtained from nonerosive reflux disease (NERD) patients and from erosive esophagitis patients. Then, the biopsies were mounted in an Ussing chamber to measure the basal transepithelial electrical resistance (TEER). The effect of mucosal exposure to an acid solution on TEER was analyzed with or without different concentrations (1, 0.3 or 1%) of SP-Gc, H-SP-Gc, or D-SP-Gc, precoated on the mucosa. Basal esophageal mucosal electrical resistance was significantly lower in erosive esophagitis than from NERD. Mucosal samples precoated with native SP-Gc (1%) significantly prevented TEER drop induced by an acidic solution in NERD, but this effect was not observed in erosive esophagitis. Topical application of D-SP-Gc showed no difference compared to native SP-Gc. However, when treated with chemically-modified SP-Gc, the protective effect observed with native SP-Gc was lost. The present study indicated that SP-Gc protects the human esophageal mucosal barrier in NERD patients. This effect is dependent on the structure but is independent of the presence of sulfate.

Independent and synergistic activity of the flavonoids of Gracilaria corticata as promising antidiabetic agents

The therapeutic approaches for Type 2 Diabetes Mellitus rely most on the usage of oral hypoglycaemic drugs. These drugs have adverse side effects and hence alternative medicines are continuously explored. The present study intends to investigate the antidiabetic potential of the flavonoids present in Gracilaria corticata. The flavonoids were isolated (FEGC) and their inhibitory activity on the carbohydrate hydrolysing enzymes such as α-amylase and α-glucosidase was analysed. The flavonoids were found to inhibit α-amylase and α-glucosidase with an IC50 value of 302 µg and 75 µg respectively. The synergistic effect of FEGC and luteolin was also investigated and the results show that both FEGC and luteolin inhibited synergistically at half their IC50 values. The observations of this study reveal that the flavonoids of G. corticata have potential antidiabetic activity and can act independently or synergistically in the management of Type 2 Diabetes Mellitus

Dual influence of terrestrial and marine historical processes on the phylogeography of the Brazilian intertidal red alga Gracilaria caudata.

In this study, we explored how past terrestrial and marine climate changes have interacted to shape the phylogeographic patterns of the intertidal red seaweed Gracilaria caudata, an economically important species exploited for agar production in the Brazilian north-east. Seven sites were sampled along the north-east tropical and south-east sub-tropical Brazilian coast. The genetic diversity and structure of G. caudata was inferred using a combination of mitochondrial (COI and cox2-3), chloroplast (rbcL) and 15 nuclear microsatellite markers. A remarkable congruence between nuclear, mitochondrial and chloroplast data revealed clear separation between the north-east (from 03° S to 08° S) and the south-east (from 20° S to 23° S) coast of Brazil. These two clades differ in their demographic histories, with signatures of recent demographic expansions in the north-east and divergent populations in the south-east, suggesting the maintenance of several refugia during the last glacial maximum due to sea-level rise and fall. The Bahia region (around 12° S) occupies an intermediate position between both clades. Microsatellites and mtDNA markers showed additional levels of genetic structure within each sampled site located south of Bahia. The separation between the two main groups in G. caudata is likely recent, probably occurring during the Quaternary glacial cycles. The genetic breaks are concordant with (i) those separating terrestrial refugia, (ii) major river outflows and (iii) frontiers between tropical and subtropical regions. Taken together with previously published eco-physiological studies that showed differences in the physiological performance of the strains from distinct locations, these results suggest that the divergent clades in G. caudata correspond to distinct ecotypes in the process of incipient speciation and thus should be considered for the management policy of this commercially important species.

Sulfated polysaccharides from the marine algae Gracilaria caudata prevent tissue damage caused by ligature-induced periodontitis.

Sulfated polysaccharides (PLS) extracted from the marine algae of the genus Gracilaria showed biological activity in different inflammatory models, except for periodontitis. Thus, this study aimed to evaluate the effectiveness of the treatment with PLS from Gracilaria caudata in ligature-induced periodontitis. 40 animals distributed into 5 groups were used (the control group (unligated), the ligated untreated group, and the ligated groups treated with 2.5, 5.0 and 10.0 mg/kg of PLS with intraperitoneal injection, respectively). After 20 days of treatment, the animals were killed and the following parameters were evaluated: Gingival Bleeding Index (GBI), Probing Pocket Depth (PPD), Myeloperoxidase (MPO) activity, Alveolar Bone Loss (ABL) for periodontal tissues; histopathological examination of gingival and liver tissues (Steatosis score); glutathione and malondialdehyde concentrations in the liver, serum levels of alanine aminotransferase and aspartate aminotransferase. The data revealed that treatment with 2.5 mg/kg of PLS showed the best anti-inflammatory effects with reduction of GBI, PPD and MPO activity, as well as oxidative stress and steatosis in liver. Our results indicated that the adjunct treatment with PLS from Gracilaria caudata could prevent the periodontal and hepatic tissue alteration caused by periodontitis.

Effects of a co-culture of marine algae and shrimp (Litopenaeus vannamei) on the growth, survival and immune response of shrimp infected with Vibrio parahaemolyticus and white spot virus (WSSV).

In aquaculture, fighting infectious diseases is a necessity. This study measured the immuno-stimulating effect of live macroalgae consumption on Litopenaeus vannamei against Vibrio parahaemolyticus and WSSV infection in two independent bioassays. Shrimps and macroalgae were cultivated in a co-culture with two species of macroalgae separately (Gracilaria vermiculophylla and Dictyota dichotoma), and later, shrimp were infected with V. parahaemolyticus. In another bioassay, shrimp and macroalgae (G. vermiculophylla, D. dichotoma and Ulva lactuca) were grown and subsequently infected with WSSV. For both bioassays, survival after 120 h was determined, the total hemocyte count (TCH) was measured and the activity of superoxide dismutase (SOD) and catalase (CAT) in tissue were measured. The results indicate that the use of macroalgae in co-culture with L. vannamei provides a nutritional benefit that achieves higher growth than the control organisms, as well as improvements of the ammonium concentration and immune response after infection with V. parahaemolyticus and WSSV. A better immune response was obtained in organisms cultured with macroalgae in both bioassays at a ratio of 1.6-1.9 for organisms infected with bacteria and 1.4 to 1.6 times for organisms infected with the virus. In turn, the enzymatic activity of SOD and CAT were higher in the treated organisms relative to the controls in both experiments.

Differentiation of haploid and diploid fertilities in Gracilaria chilensis affect ploidy ratio.

BACKGROUND: Algal isomorphic biphasic life cycles alternate between free-living diploid (tetrasporophytes) and haploid (dioicious gametophytes) phases and the hypotheses explaining their maintenance are still debated. Classic models state that conditional differentiation between phases is required for the evolutionary stability of biphasic life cycles while other authors proposed that the uneven ploidy abundances observed in the field are explained by their cytological differences in spore production. RESULTS: We monitored the state and fate of individuals of the red seaweed Gracilaria chilensis periodically for 3 years in five intertidal pools from two sites with distinct conditions. We tested for differentiation in fecundity and spore survival among the gametophyte males and females (haploids) and the tetrasporophytes (diploids). We tested for the influence of fecundity and spore survival on the observed uneven ploidy abundances in recruits. The probability of a frond becoming fecund was size-dependent, highest for the haploid males and lowest for the haploid females, with the diploids displaying intermediate probabilities. Fecund diploids released more tetraspores than carpospores released by the haploid females. Spore survival depended on ploidy and on the local density of co-habiting adult fronds. An advantage of diploid over haploid germlings was observed at very low and very high adult fronds densities. CONCLUSIONS: Neither spore production nor spore survival determined the highly variable ploidy ratio within G. chilensis recruits. This result invalidates the hypothesis of natural cytological differences in spore production as the only driver of uneven field ploidy abundances in this species. Diploid spores (carpospores) survived better than haploid spores (tetraspores), especially in locations and time periods that were associated with the occurrence of strong biotic and abiotic stressors. We hypothesise that carpospore survival is higher due to support by their haploid female progenitors passing-on nutrients and chemical compounds improving survival under stressful conditions.

Haploid females in the isomorphic biphasic life-cycle of Gracilaria chilensis excel in survival.

BACKGROUND: Conditional differentiation is one of the most fundamental drivers of biodiversity. Competitive entities (usually species) differ in environmental or ecological niche enabling them to co-exist. Conditional differentiation of haploid and diploid generations is considered to be a requirement for the evolutionary stability of isomorphic biphasic life-cycles and the cause for the natural occurrence of both phases at uneven abundances. Theoretically, stage dependent survival rates are the most efficient way to explain conditional differentiation. RESULTS: We tested for conditional differentiation in survival rates among life stages (haploid males, haploid females, and diploids) of Gracilaria chilensis, an intertidal red alga occurring along the Chilean shores. Therefore, the fate of individuals was followed periodically for 3 years in five intertidal pools and, for the first time in isomorphic red algae, a composite model of the instantaneous survival rates was applied. The results showed the survival dependency on density (both competition and Allee effects), fertility, age, size, season and location, as well as the differentiation among stages for the survival dependencies of these factors. The young haploid females survived more than the young of the other stages under Allee effects during the environmentally stressful season at the more exposed locations, and under self-thinning during the active growth season. Furthermore, fertile haploid females had a higher survival than fertile haploid males or fertile diploids. CONCLUSIONS: Here, we show a survival advantage of haploids over diploids. The haploid females probably optimize their resource management targeting structural and physiological adaptations that significantly enhance survival under harsher conditions. In a companion paper we demonstrate a fertility advantage of diploids over haploids. Together, the survival and fertility differentiation support the evolution and prevalence of biphasic life-cycles.

Nutrients, but not genetic diversity, affect Gracilaria chilensis (Rhodophyta) farming productivity and physiological responses.

In terrestrial plants, it is well known that genetic diversity can affect responses to abiotic and biotic stress and have important consequences on farming. However, very little is known about the interactive effects of genetic and environmental factors on seaweed crops. We conducted a field experiment on Gracilaria chilensis to determine the effect of heterozygosity and nutrient addition on two southern Chilean farms: Ancud and Chaica. In addition to growth rate and productivity, we measured photosynthetic responses, photosynthetic pigment concentration (chlorophyll a and phycobiliproteins), C:N ratio (C:N), and epiphytic load. Nutrient addition affected the growth rate, productivity, phycobilin, and C:N content, but not the epiphytic load. These results were independent of the heterozygosity of the strains used in the experiments. Interestingly, depending on the sampled sites, distinct photosynthetic responses (i.e., maximal quantum yield, Fv /Fm , and maximal electron transport rate, ETRmax ) to nutrient addition were observed. We propose that thallus selection over the past few decades may have led to ecological differentiation between G. chilensis from Chaica and Ancud. The lack of effect of heterozygosity on growth and physiological responses could be related to the species domestication history in which there is a limited range of genetic variation in farms. We suggest that the existing levels of heterozygosity among our thalli is not sufficient to detect any significant effect of genetic diversity on growth or productivity in Metri bay, our experimental site located close to the city of Puerto Montt, during summer under nitrogen limiting conditions.

In Vivo Evaluation of the Antioxidant Activity and Protective Action of the Seaweed Gracilaria birdiae.

The red seaweed Gracilaria birdiae (GB) is farmed and used as food in northeast Brazil. However, the economic potential of this seaweed has been explored little. To enable direct consumption and/or product diversification from GB, it is necessary to evaluate its effect in vivo. In this study, the food of mice was improved with the addition of GB. After 21 days, the consumption of seaweed reduced the weight gain and blood glucose levels in mice. In addition, it increased the trolox equivalent antioxidant capacity and glutathione reductase and catalase levels compared to those of the control group. In addition, some mice also received carbon tetrachloride (CCl4). In this case, histological, enzymatic, and antioxidant tests showed that the seaweed could protect animals from damage caused by this toxic agent. In addition, GB aqueous extract (AE) inhibited 50% of 3T3-L1 cell differentiation into adipocytes, whereas GB ethanolic extract was not effective. AE is composed mainly of sulfated polysaccharides. The results of the present study indicate that the alga GB protected the mice from CCl4-induced damage, indicating that the seaweed exhibits protective action in vivo. In addition, GB decreased the animal weight gain, which was mainly due to the action of the sulfated polysaccharides synthesized by this seaweed.

Antidiarrheal activity of a novel sulfated polysaccharide from the red seaweed Gracilaria cervicornis.

ETHNOPHARMACOLOGICAL RELEVANCE: The use of marine seaweeds as a source of natural compounds with medicinal purposes is increasing in Western countries in the last decades, becoming an important alternative in the traditional medicine of many developing countries, where diarrhea still remains a severe public health problem, with high rates of mortality and morbidity. Sulfated polysaccharides (PLS) extracted from red seaweeds can exhibit therapeutic effects for the treatment of gastrointestinal disorders. Thus, the pharmacological properties of the PLS from Gracilaria cervicornis, an endemic seaweed found in the Brazilian northeast coast, was evaluated as an alternative natural medication for diarrhea. AIM OF THE STUDY: This study aimed to evaluate the antidiarrheal activity of sulfated polysaccharides (PLS) extracted from the red seaweed G. cervicornis in Swiss mice pre-treated with castor oil or cholera toxin. MATERIALS AND METHODS: The seaweed Gracilaria cervicornis was collected at Flecheiras beach (city of Trairí, State of Ceará, Brazil) and the PLS was obtained through enzymatic extraction and administered in mice (25-30 g) before diarrhea induction with castor oil or cholera toxin. For the evaluation of the total number of fecal output and diarrheal feces, the animals were placed in cages lined with adsorbent material. The evaluation of intestinal fluid accumulation (enteropooling) on castor oil-induced diarrhea in mice occurred by dissecting the small intestine and measuring its volume. The determination of Na+/K+-ATPase activity was measured in the small intestine supernatants by colorimetry, using commercial biochemistry kits. The gastrointestinal motility was evaluated utilizing an activated charcoal as a food tracer. The intestinal fluid secretion and chloride ion concentration were evaluated in intestinal closed loops in mice with cholera toxin-induced secretory diarrhea. The binding ability of PLS with GM1 and/or cholera toxin was evaluated by an Enzyme-Linked Immunosorbent Assay (ELISA). RESULTS: The G. cervicornis PLS showed antidiarrheal effects in both acute and secretory diarrhea, reducing the total number of fecal output, diarrheic stools, intestinal fluid accumulation, and increasing small intestine Na+/K+-ATPase activity on castor oil-induced diarrhea. However, the PLS did not affect gastrointestinal motility, indicating that this compound has a different action mechanism than loperamide. In secretory diarrhea, the PLS decreased intestinal fluid secretion and small intestine chloride excretion, binding with GM1 and/or cholera toxin and blocking their attachment to the enterocyte cell surface. CONCLUSIONS: In conclusion, PLS has a significant antidiarrheal effect in acute and secretory diarrhea. Further investigation is needed towards its use as a natural medicine to treat diarrhea.