Ricardo N. Alves

In vitro bioaccessibility of the marine biotoxin okadaic acid in shellfish

Okadaic acid (OA) and their derivatives are marine toxins responsible for the human diarrhetic shellfish poisoning (DSP). To date the amount of toxins ingested in food has been considered equal to the amount of toxins available for uptake by the human body. In this study, the OA fraction released from the food matrix into the digestive fluids (bioaccessibility) was assessed using a static in vitro digestion model. Naturally contaminated mussels (Mytilus galloprovincialis) and donax clams (Donax sp.), collected from the Portuguese coast, containing OA and dinophysistoxin-3 (DTX3) were used in this study. Bioaccessibility of OA total content was 88% and 75% in mussels and donax clams, respectively. Conversion of DTX3 into its parent compound was verified during the simulated digestive process and no degradation of these toxins was found during the process. This is the first study assessing the bioaccessibility of OA-group toxins in naturally contaminated seafood. This study provides relevant new data that can improve and lead to more accurate food safety risk assessment studies concerning these toxins.

Ecophysiological responses of juvenile seabass (Dicentrarchus labrax) exposed to increased temperature and dietary methylmercury

The ecotoxicological effects of methylmercury (MeHg) exposure have been intensively described in literature. Yet, it is still unclear how marine biota will respond to the presence of MeHg under climate change, namely ocean warming. The present study aimed to investigate, for the first time, fish condition [Fultons K index (K), hepatosomatic index (HIS) and brain-to-body mass ratio (BB-ratio)] and several stress-related responses in an ecologically and commercially important fish species (Dicentrachus labrax) exposed for 28days to dietary MeHg (8.0mg kg-1 dw) and temperature increase (+4°C). Results showed significant impairments on fish condition, i.e. up to 34% decrease on K, >100% increase on HIS and 44% decrease on BB-ratio, compared to control conditions. Significant changes on tissue biochemical responses were observed in fish exposed to both stressors, acting alone or combined, evidencing the relevance of assessing possible interactions between different environmental stressors in ecotoxicological studies. For instance, muscle showed to be the least affected tissue, only revealing significant alterations in GST activity of MeHg-enriched fish. On the other hand, liver exhibited a significant induction of GST (>100%) and CAT (up to 74%) in MeHg-enriched fish, regardless of temperature exposure, as well as decreased SOD activity (19%) and increased HSP70/HSC70 content (87%) in fish exposed to warming alone. Brain showed to be affected by temperature (69% of GST inhibition and >100% of increased CAT activity), MeHg (>100% of increased CAT activity, 47% of SOD inhibition and 55% of AChE inhibition), as well as by the combination of both (GST, SOD and AChE inhibition, 17%, 48% and 53%, respectively). Hence, our data provides evidences that the toxicological aspects of MeHg ca be potentiated by warmer temperatures, thus, evidencing the need for further research combining contaminants exposure and climate change effects, to better forecast ecological impacts in the ocean of tomorrow.

Oral bioaccessibility of toxic and essential elements in raw and cooked commercial seafood species available in European markets

The oral bioaccessibility of several essential and toxic elements was investigated in raw and cooked commercially available seafood species from European markets. Bioaccessibility varied between seafood species and elements. Methylmercury bioaccessibility varied between 10 (octopus) and 60% (monkfish). Arsenic (>64%) was the toxic element showing the highest bioaccessibility. Concerning essential elements bioaccessibility in raw seafood, selenium (73%) and iodine (71%) revealed the highest percentages. The bioaccessibility of elements in steamed products increased or decreased according to species. For example, methylmercury bioaccessibility decreased significantly after steaming in all species, while zinc bioaccessibility increased in fish (tuna and plaice) but decreased in molluscs (mussel and octopus). Together with human exposure assessment and risk characterization, this study could contribute to the establishment of new maximum permissible concentrations for toxic elements in seafood by the European food safety authorities, as well as recommended intakes for essential elements.

Differential behavioural responses to venlafaxine exposure route, warming and acidification in juvenile fish (Argyrosomus regius)

Antidepressants, such as venlafaxine (VFX), which are considered emerging environmental pollutants, are increasingly more present in the marine environment, and recent evidence suggest that they might have adverse effects on fish behaviour. Furthermore, altered environmental conditions associated to climate change (e.g. warming and acidification) can also have a determinant role on fish behaviour, fitness and survival. Yet, the underlying interactions between these environmental stressors (pharmaceuticals exposure and climate change) are still far from being fully understood. The aim of this study was to assess behavioural responses (in juvenile meagre (Argyrosomus regius) exposed to VFX via water ([VFX] ~20μgL-1) and via dietary sources ([VFX] ~160μgkg-1 dry weight), as well as to increased temperature (ΔT°C=+5°C) and high CO2 levels (ΔpCO2 ~1000μatm; equivalent to ΔpH=-0.4units). Overall, VFX bioaccumulation in fish plasma was enhanced under the combination of warming and acidification. VFX triggered fish exploration, whereas fish activity and shoal cohesion were reduced. Acidification alone decreased fish exploration and shoal cohesion, and reversed fish preference to turn leftwards compared to control conditions. Such alterations were further enhanced by VFX exposure. The combination of warming and acidification also reduced shoal cohesion and loss of lateralization, regardless of VFX exposure. The distinct behaviour observed when VFX contamination, acidification and warming acted alone or in combination highlighted the need to consider the likely interactive effects of seawater warming and acidification in future research regarding the toxicological aspects of chemical contaminants.

Polycyclic aromatic hydrocarbons bioaccessibility in seafood: Culinary practices effects on dietary exposure

&NA; This work aimed to determine the effect of culinary practices on the contamination level and bioaccessibility of polycyclic aromatic hydrocarbons (PAHs) in seafood. The selected farmed seafood species (marine shrimp, clams and seaweed) were commercially available in Portugal. The mean concentrations of PAHs varied between 0.23 and 51.8 &mgr;g kg−1, with the lowest value being observed in raw shrimp and the highest in dried seaweed. The number of compounds detected in seaweed and clams (naphthalene, acenaphthene, fluorene, phenanthrene, benzo(b)fluoranthene and benzo(j)fluoranthene) were higher than in shrimp (fluorene and pyrene). Among the PAHs measured, fluorene was the predominant one. There was a significant interaction effect between species and culinary treatment (p < 0.05), thus boiled and dried seaweed samples presented the lowest and the highest levels of fluorene (0.13 and 1.8 &mgr;g kg−1), respectively. The daily intake of PAHs decreased with bioaccessibility, varying from 22% for benzo(k)fluoranthene (in raw clam) to 84% for phenanthrene (in steamed clam). According to the potency equivalent concentrations, screening values and bioaccessibility of PAHs, the consumption of marine shrimp, clam and seaweed is considered as safe for consumers. HighlightsPAHs bioaccessibility in seafood was assessed.Raw shrimp present the lowest and dried seaweed the highest content of PAH.There was a significant interaction effect between species and culinary treatment.

Will seabass (Dicentrarchus labrax) quality change in a warmer ocean

The impacts of climate change on seafood quality, safety and human health are still unknown. The present study investigated the effect of warming on fatty acids and elements content in two tissues (muscle and liver) of the relevant commercial seabass species (Dicentrarchus labrax). After exposing fish to increased seawater temperature for a period of 60days, higher saturated fatty acid (SFA) levels were observed in fish muscle (2.16% increase); whereas lower SFA levels were observed in fish liver (5.42% decrease). On the other hand, monounsaturated fatty acids (MUFA) and polyunsaturated fatty acids (PUFA) contents decreased in both muscle (1.77% and 0.39%, respectively) and liver (10.54% and 8.11%, respectively) of fish subjected to warmer conditions. Additionally, warming promoted changes in fish elemental profiles, leading to significantly higher levels of Cl in the muscle and lower levels of Rb in the liver. Overall, data showed that fatty acids and elemental contents were affected by temperature, though representing small implications to human health. Moreover, this preliminary study highlights the importance of conducting further seafood risk-benefit assessments under climate change contexts.

First approach to assess the bioaccessibility of bisphenol A in canned seafood

Human health risks due to bisphenol A (BPA) exposure through canned food consumption are an emerging safety concern worldwide. In this study, an in vitro digestion model was used to simulate human digestion and evaluate BPA bioaccessibility in canned seafood for the first time. BPA contents of canned tuna and sardine samples and their bioaccessible and non-bioaccessible fractions were determined by gas chromatography coupled to mass spectrometry (GC-MS). The 21 samples of canned tuna and sardines, all from the same producer but with different kind of sauces, showed BPA levels ranging from <1µgkg-1 (limit of quantification, LOQ) to 62µgkg-1, with variable results within and between sample groups. BPA bioaccessibility was evaluated in six positive samples, with values ranging from 80 to 99%. The results suggest that BPA bioaccessibility was slightly lower in samples with higher lipid content.

Effects of steaming on contaminants of emerging concern levels in seafood

Seafood consumption is a major route for human exposure to environmental contaminants of emerging concern (CeCs). However, toxicological information about the presence of CeCs in seafood is still insufficient, especially considering the effect of cooking procedures on contaminant levels. This study is one among a few who evaluated the effect of steaming on the levels of different CeCs (toxic elements, PFCs, PAHs, musk fragrances and UV-filters) in commercially relevant seafood in Europe, and estimate the potential risks associated with its consumption for consumers. In most cases, an increase in contaminant levels was observed after steaming, though varying according to contaminant and seafood species (e.g. iAs, perfluorobutanoate, dibenzo(ah)anthracene in Mytilus edulis, HHCB-Lactone in Solea sp., 2-Ethylhexyl salicylate in Lophius piscatorius). Furthermore, the increase in some CeCs, like Pb, MeHg, iAs, Cd and carcinogenic PAHs, in seafood after steaming reveals that adverse health effects can never be excluded, regardless contaminants concentration. However, the risk of adverse effects can vary. The drastic changes induced by steaming suggest that the effect of cooking should be integrated in food risk assessment, as well as accounted in CeCs regulations and recommendations issued by food safety authorities, in order to avoid over/underestimation of risks for consumer health.

Preliminary assessment of galaxolide bioaccessibility in raw and cooked FISH

Generally, dietary intake assessment and risk characterization are evaluated using contaminant concentration in raw fish while it is usually ingested cooked, which can cause an overestimation because one of the essential issues for risk-benefit analysis is to determine the maximum amount of a contaminant that can be released from the food matrix and be absorbed by the human body, called bioaccessibility. Moreover, despite most seafood products are cooked before consumption, risk assessment is still evaluated in raw products, strongly affecting public health guidelines. In the present study, an in vitro bioaccessibility assay was performed for Galaxolide (HHCB) in fish samples. Raw spiked hake samples were in vitro digested and aliquots of each fraction of the digestion process were analysed. HHCB was quantitatively present in the bioaccessibility fraction. The effect of fish cooking on HHCB was also evaluated in cod and mackerel samples. Results demonstrate that steaming and grilling processes lead to a loss of 50-70% HHCB in fish.

Living in a multi-stressors environment: an integrated biomarker approach to assess the ecotoxicological response of meagre (Argyrosomus regius) to venlafaxine, warming and acidification

Abstract Pharmaceuticals, such as the antidepressant venlafaxine (VFX), have been frequently detected in coastal waters and marine biota, and there is a growing body of evidence that these pollutants can be toxic to non‐target marine biota, even at low concentrations. Alongside, climate change effects (e.g. warming and acidification) can also affect marine species’ physiological fitness and, consequently, compromising their ability to cope with the presence of pollutants. Yet, information regarding interactive effects between pollutants and climate change‐related stressors is still scarce. Within this context, the present study aims to assess the differential ecotoxicological responses (antioxidant activity, heat shock response, protein degradation, endocrine disruption and neurotoxicity) of juvenile fish (Argyrosomus regius) tissues (muscle, gills, liver and brain) exposed to VFX (via water or feed), as well as to the interactive effects of warming (&Dgr;T °C = +5 °C) and acidification (&Dgr;pCO2 ˜ +1000 &mgr;atm, equivalent to &Dgr;pH = −0.4 units), using an integrated multi‐biomarker response (IBR) approach. Overall, results showed that VFX toxicity was strongly influenced by the uptake pathway, as well as by warming and acidification. More significant changes (e.g. increases surpassing 100% in lipid peroxidation, LPO, heat shock response protein content, HSP70/HSC70, and total ubiquitin content, Ub,) and higher IBR index values were observed when VFX exposure occurred via water (i.e. average IBR = 19, against 17 in VFX‐feed treatment). The co‐exposure to climate change‐related stressors either enhanced (e.g. glutathione S‐transferases activity (GST) in fish muscle was further increased by warming) or attenuated the changes elicited by VFX (e.g. vitellogenin, VTG, liver content increased with VFX feed exposure acting alone, but not when co‐exposed with acidification). Yet, increased stress severity was observed when the three stressors acted simultaneously, particularly in fish exposed to VFX via water (i.e. average IBR = 21). Hence, the distinct fish tissues responses elicited by the different scenarios emphasized the relevance of performing multi‐stressors ecotoxicological studies, as such approach enables a better estimation of the environmental hazards posed by pollutants in a changing ocean and, consequently, the development of strategies to mitigate them. HighlightsVFX toxicity was influenced by exposure route, as well as by abiotic stressors.VFX water exposure induced more severe biomarker responses than VFX feed exposure.Muscle, liver and brain biomarker responses were significantly affected by warming.Biomarker changes due to acidification were more evident in fish gills.The combination of the three stressors simultaneously increased stress severity.