Set up and validation of a method to analyse microplastics in stool and small intestine samples.

The contamination of microplastics in humans is of increasing concern. Therefore, the aim of this study was to develop effective methods to determine the concentration and types of microplastics entering human digestive system. To study levels of MPs contamination in humans, an excellent indicator are stools. Indeed, stools, and thus the digestive system, can be an excellent indicator of the level of MPs contamination in humans. Hence, objective was to find effective methods to extract, quantify and characterize microplastics in stool and small intestine samples. The samples studied were human stools and pig jejunum (which has human-like characteristics). The methods were optimized by observing extraction efficiency, compatibility by Fourier-transform infrared spectroscopy (FTIR) characterization and non-deformation of the microplastics. The steps of the procedure were: • Sampling to avoid plastic contamination • Non-aggressive chemical and enzymatic digestion • Counting and characterization The methods were optimized and validated, observing recovery and repeatability. Therefore, two simple, effective methods with high analytical performance have been developed. The MPs present in the stool and intestine samples were counted by stereoscopic microscope and characterized by FTIR, finding several types of MPs such as synthetic cellulose, polyethylene, polypropylene, polystyrene, and polyethylene terephthalate, among others.

Tracing the fate of microplastic in wastewater treatment plant: A multi-stage analysis of treatment units and sludge.

Microplastic (MP) pollution is ubiquitous in the environment presenting a global problem for both scientists and the general public. One of the major pathways of MPs entering the natural environment is through wastewater treatment plants (WWTPs). Once MPs reach the natural environment, they are posing threat to aquatic ecosystems and public health. The aim of this study is to investigate the concentration, morphology, and composition of MPs in different treatment units of a WWTP. Sampling included different points across WWTP in the water and sludge lines. Pre-treatment of the samples consists of advanced Fenton oxidation, and alkaline and enzymatic digestion followed by density separation. Once the particles were isolated, their morphology and size were studied using a stereoscopic and optical microscope followed by final confirmation with ATR-FTIR and micro-FTIR spectroscopy. Microplastic particle concentrations exhibit significant reductions as water undergoes treatment in the WWTP. For summer sampling, concentrations decreased from 351 MP/L (influent) to 35 MP/L (primary clarifier), 32 MP/L (biological reactor), and 13 MP/L (2.3 MP/L) (secondary clarifier). Similarly, winter sampling showed reductions from 403 MP/L (influent) to 159 MP/L (primary clarifier), 178 MP/L (biological reactor), and 26 MP/L (5.6 MP/L) (secondary clarifier). Removal efficiency of WWTP is high and exceeds 96%. The most abundant morphology is fibers followed by fragments and films. Polymers such as PE, synthetic cellulose, PP, PVC, PE-PP, PEEA, PA, acrylamide, and PES are widely detected in different units of WWTP. The number of MPs that are avoided from being emitted into the environment through direct water discharge was estimated to be 9.1 × 1012 MP/year. Removed MPs tend to accumulate in the sludge that is used for agricultural purposes although it should be managed as waste properly, avoiding the transition of MPs pollutants to terrestrial ecosystems adding to the number of MPs that will inevitably end up in receiving water bodies through direct WWTP effluent discharge that was set in 5.1 × 1010 MP/year in the studied WWTP.

Screening of microplastics in water and sludge lines of a drinking water treatment plant in Catalonia, Spain.

Microplastics (MPs) are emerging pollutants detected everywhere in the environment, with the potential to harm living organisms. The present study investigated the concentration, morphology, and composition of MPs, between 20 µm and 5 mm, in a drinking water treatment plant (DWTP) located close to Barcelona (Catalonia, NE Spain). The sampling included different units of the DWTP, from influent to effluent as well as sludge line. Sampling strategy, filtration, allows sampling of large volumes of water avoiding sample contamination, and during 8 h in order to increase the representativeness of MPs collected. The pre-treatment of the samples consisted of advanced oxidation with Fenton's reagent and hydrogen peroxide, followed by density separation of the particles with zinc chloride solution. Visual identification was performed with an optical and stereoscopic microscope with final Fourier-transform infrared spectroscopic (FTIR) confirmation. MPs were found in all DWTP samples, with concentrations from 4.23 ± 1.26 MPs/L to 0.075 ± 0.019 MPs/L in the influent and effluent of the plant, respectively. The overall removal efficiency of the plant was 98.3%. The most dominant morphology was fibers followed by fragments and films. Twenty-two different polymer types were identified and synthetic cellulose, polyester, polyamide, polypropylene, polyethylene, polyurethane, and polyacrylonitrile were the most common. Although MPs could be incorporated from the distribution network, MPs intake from drinking water from this DWTP was not an important route compared to fish and seafood ingestion.

Levels of microplastics and their characteristics in molluscs from North-West Mediterranean Sea: Human intake.

Microplastics (MPs) are accessible for organisms with active filter feeding strategies, as are many marine molluscs, which live attached or semi-buried in sediments. In the present study, MPs (from 0.02 to 5 mm) concentration, morphology, and composition were determined in consumed mollusc species of the Catalan coast (NW Mediterranean Sea). Microplastic concentrations, morphologic characteristics and composition were studied according to species, catchment zones and depuration condition. Finally, human intake of MPs through molluscs' consumption was determined. >2300 individuals were analysed, being 1460 MPs extracted and their size, and polymeric composition registered. Big oysters and mussels showed the highest MPs concentration by individual, with levels of 22.8 ± 14.4 and 18.6 ± 23.0 MPs/individual, respectively. Mean annual MPs (≥20 µm) consumption for adult population was estimated in 8103 MPs/year, with a 95th percentile of 19,418 MPs/year. It suggests that the consumption of molluscs is an important route of MPs exposure for the Catalan population.

Microplastics levels, size, morphology and composition in marine water, sediments and sand beaches. Case study of Tarragona coast (western Mediterranean).

Mediterranean Sea has been proposed as the sixth greatest accumulation zone for marine litter and the most affected regarding to microplastics (MPs). Tarragona (Catalonia, NE Spain) coastal region suffers high pressure due to urbanization, tourism, industrial harbour and petrochemical/plastic industries. The present study aims to quantify and characterize in size, morphology and composition the MPs present in sandy beaches, marine sediments, and surface seawaters of Tarragona coastal region. MPs mean abundance were 1.30 items/m3 in surface seawaters, 32.4 items/kg in marine sediments, and 10.7 items/kg in sandy beaches. Polyester fibres were dominant MPs in bottom sediments and seawater meanwhile polyethylene and polypropylene fragments were the main MPs in beaches. The fibres balls associated with bottom sediments, organic matter and plankton were abundant, masking the real quantity of fibres in each reservoir. The abundance by volume of seawater MPs was higher to those found in oceanic areas and similar to other areas of Mediterranean Sea, corroborating that Western Mediterranean Sea as a region of MPs accumulation. MPs composition and abundance suggested the input of numerous land-base-sources, WWTP (wastewater treatment plants) effluents discharges, and emissaries as the most important. Marine MPs pollution were studied from an integrative point of view, that includes superficial sea water, sand from beaches and sediments. The dynamics of MPs in Tarragona coast were characterized by seawater as the media that receive and facilitate dispersion and fragmentation. The shoreline acts as an intermediate reservoir with constant weathering and active exchange with seawater surface and the sediments acts as a significant sink for medium MPs sizes. It is necessary to develop protocols and guidelines for MPs analysis to obtain harmonized and comparable results.

Performance of Chlorella Vulgaris Exposed to Heavy Metal Mixtures: Linking Measured Endpoints and Mechanisms.

Microalgae growth inhibition assays are candidates for referent ecotoxicology as a fundamental part of the strategy to reduce the use of fish and other animal models in aquatic toxicology. In the present work, the performance of Chlorella vulgaris exposed to heavy metals following standardized growth and photosynthesis inhibition assays was assessed in two different scenarios: (1) dilutions of single heavy metals and (2) an artificial mixture of heavy metals at similar levels as those found in natural rivers. Chemical speciation of heavy metals was estimated with Visual MINTEQ software; free heavy metal ion concentrations were used as input data, together with microalgae growth and photosynthesis inhibition, to compare different effects and explain possible toxicity mechanisms. The final goal was to assess the suitability of the ecotoxicological test based on the growth and photosynthesis inhibition of microalgae cultures, supported by mathematic models for regulatory and decision-making purposes. The C. vulgaris algae growth inhibition test was more sensitive for As, Zn, and Pb exposure whereas the photosynthesis inhibition test was more sensitive for Cu and Ni exposure. The effects on growth and photosynthesis were not related. C. vulgaris evidenced the formation of mucilaginous aggregations at lower copper concentrations. We found that the toxicity of a given heavy metal is not only determined by its chemical speciation; other chemical compounds (as nutrient loads) and biological interactions play an important role in the final toxicity. Predictive mixture effect models tend to overestimate the effects of metal mixtures in C. vulgaris for both growth and photosynthesis inhibition tests. Growth and photosynthesis inhibition tests give complementary information, and both are a fast, cheap, and sensitive alternative to animal testing. More research is needed to solve the challenge of complex pollutant mixtures as they are present in natural environments, where microalgae-based assays can be suitable monitoring tools for pollution management and regulatory purposes.

Microplastics and metal burdens in freshwater Tilapia (Oreochromis niloticus) of a metropolitan reservoir in Central Mexico: Potential threats for human health.

In the present study, microplastics (MPs) and metal concentrations were studied in the widely consumed tilapia (Oreochromis niloticus) fishes (n = 15) collected from a metropolitan reservoir of the Atoyac River basin, Mexico. Nearly 139 fibers were extracted from the gastrointestinal tracts and assessed using optical microscopy to evaluate their physical characteristics. The colour distribution of the fibers was mainly black (40%), blue (19%), red and white (14%). SEM images represented the surface morphology, while the elemental composition of the fibers was studied using EDX spectra. Polymer characterization using µFTIR aided in confirming the fibers as plastics (polyamide, polyester, and synthetic cellulose) and non-plastics (natural cellulose). Henceforth, ∼33% of the fibers, provisionally thought to be plastics, were natural fibers. The total metal concentrations were higher in the liver (259.24 mg kg-1) than the muscle (122.56 mg kg-1) due to diverse metabolic functions in the hepatic tissues. Human health risk assessment in terms of Hazard Index (HI) presented Pb and Zn values above unity in both adults and children, prompting regulatory measures. Statistical tests between MPs and fish biometry did not present any substantial correlations. The present study also affirmed that the presence of MPs and metals in fishes of a highly contaminated region is not only governed by their bioavailabilities, but also on the physiological characteristics of the individual organism.

Performance of Raphidocelis subcapitata exposed to heavy metal mixtures.

Microalgae growth inhibition assays are candidates for referent ecotoxicological assays, and are a fundamental part in the strategy to reduce the use of fish and other animal models in aquatic toxicology. In the present work, the performance of Raphidocelis subcapitata exposed to heavy metals following standardized growth inhibition assays has been assessed in three different scenarios: 1) dilutions of single heavy metals, 2) artificial mixture of heavy metals at similar levels than those found in natural rivers and, 3) natural samples containing known mixtures of contaminants (heavy metals). Chemical speciation of heavy metals has been estimated with Eh-pH diagram and Visual MINTEQ software; heavy metal and free heavy metal ion concentrations were used as input data, together with microalgae growth inhibition, for Dr. Fit software. The final goal was to assess the suitability of the ecotoxicological test based on the growth inhibition of microalgae cultures, and the mathematic models based on these results, for regulatory and decision-making purposes. The toxicity of a given heavy metal is not only determined by its chemical speciation; other chemical and biological interaction play an important role in the final toxicity. Raphidocelis subcapitata 48h-h-EC50 for tested heavy metals (especially Cu and Zn) were in agreement with previous studies, when ion metal bioavailability was assumed to be 100%. Nevertheless, the calculated growth inhibition was not in agreement with the obtained inhibition when exposed to the artificial mixture of heavy metals or the natural sample. Interactions between heavy metal ions and the compounds of the culture media and/or the natural sample determine heavy metal bioavailability, and eventually their toxicity. More research is needed for facing the challenge posed by pollutant mixtures as they are present in natural environments, and make microalgae-based assays suitable for pollution management and regulatory purposes.