Ultrasound-assisted extraction as an easy-to-perform analytical methodology for monitoring ibuprofen and its main metabolites in mussels.

Non-steroidal anti-inflammatory drugs (NSAIDs) have been reported to be the main pharmaceutical class accumulated in seafood. Among them, ibuprofen (IBU) is of special concern as it is used worldwide to treat common pain, does not require a medical prescription, it is often taken in a high daily dose, and has been reported to cause potential adverse effects on aquatic organisms. IBU is highly transformed into hydroxy- and carboxy-metabolites and/or degradation products generated not only after its administration but also during wastewater treatment or in the environment. These compounds can be present in the environment at higher concentrations than IBU and present higher toxicity. In this work, a low-cost and affordable routine analytical method was developed and validated for the first-time determination of IBU and its main metabolites in mussels. The method is based on ultrasound-assisted extraction (UAE), clean-up by dispersive solid-phase extraction (d-SPE) and analytical determination by liquid chromatography-tandem mass spectrometry. Box-Behnken experimental design was used for method optimisation to better evaluate the influence and interactions of UAE and d-SPE variables. Extraction recoveries were in the range from 81 to 115%. Precision, expressed as relative standard deviation, was lower than 7%. Method detection limits were in the range from 0.1 to 1.9 ng g-1 dry weight. The method was successfully applied to wild mussels.

Influence of Particle Size on Ecotoxicity of Low-Density Polyethylene Microplastics, with and without Adsorbed Benzo-a-Pyrene, in Clam Scrobicularia plana.

This study investigated the ecotoxicological effects of differently sized (4-6 µm and 20-25 µm) low-density polyethylene (LDPE) microplastics (MPs), with and without adsorbed benzo-a-pyrene (BaP), in clam Scrobicularia plana. Biomarkers of oxidative stress (superoxide dismutase-SOD; catalase-CAT), biotransformation (glutathione-S-transferases-GST), oxidative damage (lipid peroxidation-LPO) and neurotoxicity (acetylcholinesterase-AChE) were analysed in gills and digestive glands at different time intervals for a total of 14 days of exposure. In order to have a better impact perspective of these contaminants, an integrated biomarker response index (IBR) and Health Index were applied. Biomarker alterations are apparently more related to smaller sized (4-6 µm) MPs in gills and to virgin LDPE MPs in the digestive gland according to IBR results, while the digestive gland was more affected by these MPs according to the health index.

Cytotoxic responses of the anticancer drug cyclophosphamide in the mussel Mytilus galloprovincialis and comparative sensitivity with human cells lines.

The rise of cancer cases worldwide led to an increase in production and consumption of anticancer drugs, that ultimately end up in the marine environment and are accumulated in aquatic organisms. Cyclophosphamide (CP) is a cytotoxic alkylating agent frequently prescribed in cancer treatments. This study assess ecotoxicological effects of CP on mussels Mytilus galloprovincialis, through in vivo and ex vivo approaches and compares the sensitivity of mussel haemocytes with well-established human cell lines (RPE and HeLa). Mussels were exposed in vivo to CP (1000 ng L-1) and several biomarkers analysed in gills and digestive glands namely neurotoxicity (AChE activity), oxidative stress (GPx activity), biotransformation (GST activity), lipid peroxidation (LPO) and apoptosis (caspase activity), whereas genotoxicity was determined in mussels' haemocytes. Cytotoxicity was also assessed in haemocytes (in vivo and ex vivo) and human cell lines (in vitro) exposed to a range of CP concentrations (50, 100, 250, 500 and 1000 ng L-1) over 24 h, via neutral red assay. In in vivo exposure, detoxification of CP did not efficiently occur in the gills while in digestive glands GPx and GST activities were induced, jointly with a decrease in lipid peroxidation, indicating a potential outcome of the protective antioxidant mechanisms, whereas no apoptosis was noted. Moreover, cytotoxicity and DNA damage were detected in haemocytes. The ex vivo exposure haemocytes to CP caused cytotoxicity (from 100 ng L-1), whereas no effects occurred in human cell lines. This suggests that, at relevant environmental concentrations, CP cause subtle and irreversible impacts on M. galloprovincialis.

Ecotoxicological assessment of chemotherapeutic agents using toxicity tests with embryos of Mellita quinquiesperforata.

The consumption of anticancer agents has increased in the recent decades, and these substances may be present in sewage. Consequently, they may reach the environment when sanitation infrastructure is ineffective. This study evaluated the toxicity of three anticancer agents-Tamoxifen (TAM), Cisplatin (CisPt), and Cyclophosphamide (CP)-on the development of embryos of the sand-dollar Mellita quinquiesperforata. Adult individuals were collected in sandy beaches, and gametes were obtained. Freshly-fertilized eggs were exposed to increasing sets of concentrations of each compound, and the effective concentrations needed to cause a 50% effect in the organisms (EC50) were calculated. The three compounds were toxic, and their EC50 values were 16.78 ± 2.42 ng·L-1 (TAM), 27.20 ± 38.26 ng·L-1 (CisPt), and 101.82 ± 70.96 ng·L-1 (CP). There is no information on the environmental levels of these compounds in Brazil, but as they were already detected in ng·L-1 levels worldwide, it can be expected that these substances pose environmental risks to the marine biota.

Effects of mixtures of anticancer drugs in the benthic polychaete Nereis diversicolor.

The increasing consumption of anticancer drugs through single and/or combinatory chemotherapy worldwide raised concern regarding their toxicity burden in coastal zones. The toxicity of a mixture of three compounds involving the drugs cisplatin (CisPt), cyclophosphamide (CP) and tamoxifen (TAM) was determined on the marine polychaete Nereis diversicolor exposed to an increasing range of their concentrations, respectively: Mix A: 0.1 + 10 + 0.1 ng L-1; Mix B: 10 + 100 + 10 ng L-1; Mix C: 100 + 500 + 25 ng L-1; Mix D: 100 + 1000 + 100 ng L-1. Different endpoints were assessed, including disturbance in the burrowing behaviour, neurotoxicity (acetylcholinesterase - AChE activity), antioxidant enzymes (superoxide dismutase - SOD; catalase - CAT; selenium-dependent glutathione peroxidase - Se-GPx and total glutathione peroxidases T-GPx activities), biotransformation metabolism (glutathione-S-transferases - GST), lipid peroxidation (LPO) and genotoxicity (DNA damage). Biological effects of the mixtures of anticancer compounds on N. diversicolor were compared with previous studies about effects on the same biological model under single-drug exposure conducted with the same molecules. Regarding SOD activity, TAM showed an antagonist effect over CisPt and CP in mixtures C and D. In Mix D, there was a synergistic effect of TAM and CisPt that inhibited CAT activity and an additive interaction of CisPt and CP on the Phase II biotransformation enzyme. Drugs in Mix A also suppressed polychaetes' GST activity, although different from the respective single-drug responses, besides able to induce T-GPx activity, that was not sufficient to avoid oxidative damage and mid-grade DNA damage. Due to the absence of burrowing impairment in Mix A, mechanisms involved in neurotoxicity were other than the one driven by AChE alterations. At the intermediary concentrations (Mix B and C), only LPO occurred. Data from drugs individually may not predict the risks provided by mixtures.

Using an integrated approach to assess the sediment quality of an estuary from the semi-arid coast of Brazil.

The Jundiaí-Potengi Estuary (JPE) on the semi-arid coast of Brazil is influenced by multiple sources of pollution. Sediment quality at 10 JPE sites was evaluated through an integrated approach. Rainy and dry seasons were considered. Collected sediments were analyzed for texture, metal, nitrogen, phosphorus concentrations, and toxicity to invertebrates. Geochemical and ecotoxicological data were integrated using qualitative approaches and multivariate techniques. We observed decreased sediment quality in both seasons, particularly in the mid-estuary. In the dry season, the contamination-toxicity relationship was clearer, as hydrological conditions favor contaminant retention within the estuary. Rainy season conditions were found to be worse, since stormwater drainage from agricultural and urban areas carries the contamination into the estuary. Because of the contamination sources and dissolved and particle-bound metal transport, contamination and toxicity did not correlate as clearly in the rainy season. The results suggest that unmeasured contaminants are contributing to JPE sediment degradation.