Nanoparticles of Lignins and Saccharides from Fishery Wastes as Sustainable UV-Shielding, Antioxidant, and Antimicrobial Biofillers.

Lignin nanoparticles containing saccharides from fishery wastes were prepared as sustainable biofillers for advanced materials. Organosolv lignin and Kraft lignin were used as polyphenol components in association with chitosan and chitooligosaccharides. The chemophysical and biological activities of lignin/saccharide nanoparticles, such as UV-shielding, antioxidant, and antimicrobial activities, were found to be dependent on both molecular weight and deacetylation degree of saccharides, with the best performance being obtained in the presence of low-molecular-weight and highly deacetylated chitooligosaccharides. In addition, chitooligosaccharides showed a synergistic antioxidant effect with lignins, associated with antimicrobial activity against Escherichia coli (Gram-negative) and Staphylococcus aureus (Gram-positive).

Synthetic control over polymorph formation in the d-band semiconductor system FeS2.

Pyrite, also known as fool's gold is the thermodynamic stable polymorph of FeS2. It is widely considered as a promising d-band semiconductor for various applications due to its intriguing physical properties. Marcasite is the other naturally occurring polymorph of FeS2. Measurements on natural crystals have shown that it has similarly promising electronic, mechanical, and optical properties as pyrite. However, it has been only scarcely investigated so far, because the laboratory-based synthesis of phase-pure samples or high quality marcasite single crystal has been a challenge until now. Here, we report the targeted phase formation via hydrothermal synthesis of marcasite and pyrite. The formation condition and phase purity of the FeS2 polymorphs are systematically studied in the form of a comprehensive synthesis map. We, furthermore, report on a detailed analysis of marcasite single crystal growth by a space-separated hydrothermal synthesis. We observe that single phase product of marcasite forms only on the surface under the involvement of H2S and sulphur vapor. The availability of high-quality crystals of marcasite allows us to measure the fundamental physical properties, including an allowed direct optical bandgap of 0.76 eV, temperature independent diamagnetism, an electronic transport gap of 0.11 eV, and a room-temperature carrier concentration of 4.14 × 1018 cm-3. X-ray absorption/emission spectroscopy are employed to measure the band gap of the two FeS2 phases. We find marcasite has a band gap of 0.73 eV, while pyrite has a band gap of 0.87 eV. Our results indicate that marcasite - that is now synthetically available in a straightforward fashion - is as equally promising as pyrite as candidate for various semiconductor applications based on earth abundant elements.

Methodological Approaches for Monitoring Five Major Food Safety Hazards Affecting Food Production in the Galicia-Northern Portugal Euroregion.

The agri-food industry has historically determined the socioeconomic characteristics of Galicia and Northern Portugal, and it was recently identified as an area for collaboration in the Euroregion. In particular, there is a need for action to help to ensure the provision of safe and healthy foods by taking advantage of key enabling technologies. The goals of the FOODSENS project are aligned with this major objective, specifically with the development of biosensors able to monitor hazards relevant to the safety of food produced in the Euroregion. The present review addresses the state of the art of analytical methodologies and techniques-whether commercially available or in various stages of development-for monitoring food hazards, such as harmful algal blooms, mycotoxins, Listeria monocytogenes, allergens, and polycyclic aromatic hydrocarbons. We discuss the pros and cons of these methodologies and techniques and address lines of research for point-of-care detection. Accordingly, the development of miniaturized automated monitoring strategies is considered a priority in terms of health and economic interest, with a significant impact in several areas, such as food safety, water quality, pollution control, and public health. Finally, we present potential market opportunities that could result from the availability of rapid and reliable commercial methodologies.

Detoxification of paralytic shellfish poisoning toxins in naturally contaminated mussels, clams and scallops by an industrial procedure.

Paralytic shellfish poisoning (PSP) episodes cause important economic impacts due to closure of shellfish production areas in order to protect human health. These closures, if are frequent and persistent, can seriously affect shellfish producers and the seafood industry, among others. In this study, we have developed an alternative processing method for bivalves with PSP content above the legal limit, which allows reducing toxicity to acceptable levels. A modification of the PSP detoxifying procedure stablished by Decision 96/77/EC of the European Union in Acanthocardia tuberculata, was developed and implemented for PSP elimination in other bivalves species. The procedure was applied to 6 batches of mussels, 2 batches of clams and 2 batches of scallops, achieving detoxification rates of around 85%. A viable industrial protocol which allows the transformation of a product at risk into a safe product was developed. Although a significant reduction was obtained, in a sample circa 9000 µg STX diHCl equiv/kg, the final toxin level in these highly toxic mussels did not fall below the European limit. The processing protocol described may be applied efficiently to mussels, clams and scallops and it may be a major solution to counteract the closure of shellfish harvesting areas, especially if persistent.

Occurrence of Tetrodotoxin in Bivalves and Gastropods from Harvesting Areas and Other Natural Spaces in Spain.

Tetrodotoxin (TTX) is a potent neurotoxin that is receiving increasing interest in the European Union because it has been found in different fishery products (fish, bivalves and gastropods) captured in European waters. Since available information is scarce, further analytical data regarding the incidence of this toxin in European fishery products is needed in order to perform an appropriate risk assessment devoted to protecting consumers' health. Hence, samples of bivalves and gastropods were collected at different points of the Spanish coast and analyzed by high-performance hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) to evaluate the presence of TTX. None of the analyzed samples showed TTX above an internal threshold of 10 µg/kg or even showed a peak under it. Our results on TTX occurrence obtained in bivalve molluscs and gastropods did not show, at least in the studied areas, a risk for public health. However, taking into account previous positive results obtained by other research groups, and since we did not detect TTX in our samples, a more completed study increasing sampling frequency is needed to ensure proper risk evaluation towards the food safety of these products.

Tetrodotoxin, an Extremely Potent Marine Neurotoxin: Distribution, Toxicity, Origin and Therapeutical Uses.

Tetrodotoxin (TTX) is a potent neurotoxin responsible for many human intoxications and fatalities each year. The origin of TTX is unknown, but in the pufferfish, it seems to be produced by endosymbiotic bacteria that often seem to be passed down the food chain. The ingestion of contaminated pufferfish, considered the most delicious fish in Japan, is the usual route of toxicity. This neurotoxin, reported as a threat to human health in Asian countries, has spread to the Pacific and Mediterranean, due to the increase of temperature waters worldwide. TTX, for which there is no known antidote, inhibits sodium channel producing heart failure in many cases and consequently death. In Japan, a regulatory limit of 2 mg eq TTX/kg was established, although the restaurant preparation of "fugu" is strictly controlled by law and only chefs qualified are allowed to prepare the fish. Due to its paralysis effect, this neurotoxin could be used in the medical field as an analgesic to treat some cancer pains.

Occurrence of lipophilic marine toxins in shellfish from Galicia (NW of Spain) and synergies among them.

Lipophilic marine toxins pose a serious threat for consumers and an enormous economic problem for shellfish producers. Synergistic interaction among toxins may play an important role in the toxicity of shellfish and consequently in human intoxications. In order to study the toxic profile of molluscs, sampled during toxic episodes occurring in different locations in Galicia in 2014, shellfish were analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS), the official method for the detection of lipophilic toxins. The performance of this procedure was demonstrated to be fit for purpose and was validated in house following European guidelines. The vast majority of toxins present in shellfish belonged to the okadaic acid (OA) group and some samples from a particular area contained yessotoxin (YTX). Since these toxins occur very often with other lipophilic toxins, we evaluated the potential interactions among them. A human neuroblastoma cell line was used to study the possible synergies of OA with other lipophilic toxins. Results show that combination of OA with dinophysistoxin 2 (DTX2) or YTX enhances the toxicity triggered by OA, decreasing cell viability and cell proliferation, depending on the toxin concentration and incubation time. The effects of other lipophilic toxins as 13-desmethyl Spirolide C were also evaluated in vitro.

Cytotoxic activity of extracts of marine sponges from NW Spain on a neuroblastoma cell line.

Six species of marine sponges collected at intertidal and sublittoral sites of the coast of Galicia (NW Spain) were screened for potential cytotoxic properties on Neuroblastoma BE(2)-M17 cell line. Exposure to Halichondria panicea, Pachymatisma johnstonia, Ophlitaspongia seriata and Haliclona sp. aqueous extracts strongly affected cell appearance, inducing loss of neuron-like morphology and the formation of clumps. Extracts from these species also caused significant rates of cell detachment and decrease of mitochondrial membrane potential. Incubation with P. johnstonia, O. seriata and Suberites massa extracts also decreased the rate of cell proliferation. The increase of incubation time enhanced propidium iodide uptake by neuroblastoma cells. Toxic responses triggered by sponge extracts are compatible with apoptotic phenomena in neuroblastoma cells, even though increasing propidium uptake at long periods of exposure might indicate the induction of secondary necrosis. The cytotoxic properties of the tested extracts suggest the presence of compounds with potential pharmacological or biotechnological applications in the screened sponge species.

Comparison between a TaqMan polymerase chain reaction assay and a culture method for ctx-positive Vibrio cholerae detection.

The main objective of the present work was to evaluate a real-time polymerase chain reaction (PCR) method to detect toxigenic Vibrio cholerae in Pangasius hypophthalmus, a freshwater fish cultured mainly in South East Asia. A FDA traditional culture method and a real-time PCR method of the ctx gene were used for detection of V. cholerae in spiked samples of pangasius fish. After an overnight enrichment of samples at 37 degrees C in alkaline peptone water, 2 cfu/25 g of fish was detected with both methods. Although both methods were very sensitive, obtaining results with culture methods may take several days, while real-time PCR takes only a few hours. Furthermore, with traditional methods, complementary techniques such as serotyping, although not available for all serogroups, are needed to identify toxigenic V. cholerae. However, with real-time PCR, toxigenic serogroups are detected in only one step after overnight enrichment.

Comparative evaluation of enzyme-linked immunoassay and reference methods for the detection of shellfish hydrophilic toxins in several presentations of seafood.

A comparative study was conducted to determine the feasibility of enzyme-linked immunosorbent assays (ELISAs) for the detection of amnesic shellfish poisoning (ASP) and paralytic shellfish poisoning (PSP) toxins in nine naturally contaminated species in fresh, frozen, boiled and canned fish and shellfish. PSP and ASP were analyzed in 138 shellfish samples (mussels, clams, barnacles, razor shells, scallops and cockles) and anchovies by mouse bioassay (MBA) and high performance liquid chromatography with ultraviolet detection (HPLC-UV), respectively. Results were compared with toxin concentrations obtained using two commercial competitive ELISAs, saxitoxin and ASP kits. Immunoassays were able to quantify toxins in different matrices showing excellent Pearson's correlation coefficients (r = 0.974 for saxitoxin ELISA and r = 0.973 for ASP ELISA) and to detect PSP and ASP with a lower limit of detection (LOD), namely, 50 microg saxitoxin equivalent/kg shellfish meat for PSP and 60 microg/kg domoic acid in shellfish flesh for ASP, than the reference methods (350 microg saxitoxin equivalent/kg shellfish meat and 1.6 mg/kg domoic acid in shellfish flesh, respectively). These results suggest that the ELISA method could be used as screening systems in a variety of species without matrix interference.

Decrease of marine toxin content in bivalves by industrial processes.

Harmful algal blooms cause important economical losses due to the accumulation of toxins in shellfish. Natural detoxification occurs but this mechanism is very slow in most cases. The achievement of a method for the rapid detoxification of commercial bivalves would be very interesting for the shellfish harvesting sector in order to diminish economical losses due to harvesting areas closure. In this work, four different methods easily applicable in the food industry (freezing, evisceration, ozonization and thermal processing) were studied to gain the detoxification of four species of bivalves (mussels, scallops, clams and cockles) contaminated with the three main types of toxins (ASP, DSP, PSP). Results show that for ASP a significant decrease of the toxin levels below the legal limit (20 microg/g) is achieved by using hepatopancreas ablation or combination of simple steps (evisceration and/or thermal processing/and or freezing). In our hands, PSP toxin levels are sharply decreased under the limit of detection (35 microg STX eq/100g) after a thermal processing, inducing percentages of detoxification higher than 50%. The effect of freezing on the levels of PSP is very dependent on the matrix studied. DSP toxins are not significantly reduced with none of these methods.

Migration of BADGE (bisphenol A diglycidyl-ether) and BFDGE (bisphenol F diglycidyl-ether) in canned seafood.

Migration of potentially toxic materials used for the lining of commercial can goods remains an important issue, especially with respect to certain types of processed foods. Seafood is one type where more information is needed with respect to other ingredients used for adding value to fishery products. Most cans are internally coated with starters of resins such as bisphenol A diglycidyl-ether (BADGE) and bisphenol F diglycidyl-ether (BFDGE), both considered as toxic compounds. Several seafood products, sardines, tuna fish, mackerel, mussels, cod and mackerel eggs, were manufactured in different conditions changing covering sauce, time and temperature of storage and heat-treated for sterilization in cans. Migration kinetics of BADGE and BFDGE from varnish into canned products were evaluated by HPLC in 70 samples after 6, 12 or 18 months of storage. Results showed that there is no migration of BADGE in tuna fish, sardines, mussels or cod. However, migration of BFDGE occurs in all species, in a storage time-dependent way and content of fat, although migration of these compounds is not affected by sterilization conditions. All samples analyzed presented values lower than 9 mg BADGE/kg net product without exceeding European limits. However, concerning BFDGE migration, European legislation does not allow the use and/or the presence of BFDGE. Main migration takes place in mackerel reaching the highest values, 0.74 mg BFDGE/kg and 0.34 mg BADGE/kg net product, in red pepper sauce.

Cytotoxic effects of BADGE (bisphenol A diglycidyl ether) and BFDGE (bisphenol F diglycidyl ether) on Caco-2 cells in vitro.

Bisphenol A diglycidyl ether (BADGE) and bisphenol F diglycidyl ether (BFDGE) are used as starting substances for the manufacturing of epoxy resins used in internal can coatings. They are obtained by a condensation reaction between epichlorohydrin with bisphenol A and bisphenol F, respectively. These potential endocrine disrupting chemicals are able to enter the food chain and to reach the intestinal epithelium, causing structural and functional damages. The human colorectal adenocarcinoma cell line Caco-2 is a widely used in vitro model of the intestinal cells. The aim of this study was to characterize BADGE and BFDGE toxicity in Caco-2 cells, in particular, at the cellular and molecular level. Using several approaches, we characterized BADGE- and BFDGE-induced cell toxicity in Caco-2 cells. The treatment was done using different concentrations up to cytotoxic doses and different times of exposure to the agents. We evaluated the effect of these compounds on cell morphology, cell detachment, cell proliferation, F-actin disruption and plasma membrane integrity. Both compounds are able to induce morphological changes, cell detachment from the substratum and to inhibit cell proliferation, being these effects time and dose-dependent. Moreover, BADGE and BFDGE induce F-actin depolymerization, this effect is very potent at 24 h of incubation with the agents and a complete F-actin disruption can be observed at 200 microM BADGE or BFDGE. In addition, cell integrity is not damaged, since neither propidium iodide uptake nor LDH release takes place in Caco-2 cells exposed to high doses of these agents for 24 h.

Collapse of mitochondrial membrane potential and caspases activation are early events in okadaic acid-treated Caco-2 cells.

Diarrhetic Shellfish Poisoning (DSP) results from the consumption of shellfish contaminated with okadaic acid (OA) or one of the dinophysistoxins (DTX). It has been reported that this toxin induces apoptosis in several cell models, but the molecular events involved in this process have not been clarified. In this report we studied intracellular signals induced by OA in Caco-2 cells: mitochondrial membrane potential, F-actin depolymerization, caspases activation, cell proliferation and cell membrane integrity. Results indicate that caspases-8 and -9 increased their activity after 30 min of OA treatment according to their role as initiator caspases. In contrast, activation of the downstream caspase-3 is a later event in the execution phase of apoptosis. Mitochondrial membrane potential changes are detected at 30 min of OA exposure indicating that this is an early response in the apoptotic cascade. F-actin depolymerization occurs after 24h of incubation with OA and this effect is significant at low doses of the toxin. LDH is released into the culture medium, although there is not PI uptake, indicative of a significant cell death in addition to apoptosis. Moreover, OA led to a dose- and time-dependent decrease in cellular proliferation.

Effect of okadaic acid on integrins and structural proteins in BE(2)-M17 cells.

Okadaic acid (OA), an algal toxin, is known to induce Diarrhetic Shellfish Poisoning and apoptosis in a variety of cell lines. One of the main targets of OA is the actin cytoskeleton which can be modulated by integrins and other structural proteins. In this paper we studied the role of these proteins and skeletal structures on OA-induced apoptosis in neuroblastoma cells. Results show that beta1 integrin and vinculin are down-regulated when cells were exposed to OA. We observed an interaction between talin and beta1 integrin that is impaired in OA treated cells.

Diagnóstico de la hidatidosis mediante la prueba de doble difusión arco 5 en ciudadanos de la clase 1964 de la Provincia del Chubut / Diagnosis of hydatidosis by means of double dyfusion test arc 5 in students of the 1964 class of the Province of Chubut

Se determinó el número de portadores de quistes hidatídicos, y su procedencia, en una muestra selecionada de ciudadanos próximos a ser incorporados al servicio militar en la Provincia del Chubut. El examen de los sueros mediante la prueba de doble difusión arco 5 permitió establecer que el 2.27 por ciento de ellos se encontraba afectado por esta patología. Los hallazgos demuestran la unidad de la encuestas inmunodiagnósticas para la detección precoz de casos y la vigilancia epidemiológica de los programas de control

Diagnóstico de la hidatidosis mediante la prueba de doble difusión arco 5 en ciudadanos de la clase 1964 de la Provincia del Chubut / Diagnosis of hydatidosis by means of double dyfusion test arc 5 in students of the 1964 class of the Province of Chubut

Se determinó el número de portadores de quistes hidatídicos, y su procedencia, en una muestra selecionada de ciudadanos próximos a ser incorporados al servicio militar en la Provincia del Chubut. El examen de los sueros mediante la prueba de doble difusión arco 5 permitió establecer que el 2.27 por ciento de ellos se encontraba afectado por esta patología. Los hallazgos demuestran la unidad de la encuestas inmunodiagnósticas para la detección precoz de casos y la vigilancia epidemiológica de los programas de control (AU)