How many microplastics do you need to (sub)sample?

Analysis of microplastics in the environment requires polymer characterization as a confirmation step for suspected microplastic particles found in a sample. Material characterization is costly and can take a long time per particle. When microplastic particle counts are high, many researchers cannot characterize every particle in their sample due to time or monetary constraints. Moreover, characterizing every particle in samples with high plastic particle counts is unnecessary for describing the sample properties. We propose an a priori approach to determine the number of suspected microplastic particles in a sample that should be randomly subsampled for characterization to accurately assess the polymer distribution in the environmental sample. The proposed equation is well-founded in statistics literature and was validated using published microplastic data and simulations for typical microplastic subsampling routines. We report values from the whole equation but also derive a simple way to calculate the necessary particle count for samples or subsamples by taking the error to the power of negative two. Assuming an error of 0.05 (5 %) with a confidence interval of 95 %, an unknown expected proportion, and a sample with many particles (> 100k), the minimum number of particles in a subsample should be 386 particles to accurately characterize the polymer distribution of the sample, given the particles are randomly characterized from the full population of suspected particles. Extending this equation to simultaneously estimate polymer, color, size, and morphology distributions reveals more particles (620) would be needed in the subsample to achieve the same high absolute error threshold for all properties. The above proposal for minimum subsample size also applies to the minimum count that should be present in samples to accurately characterize particle type presence and diversity in a given environmental compartment.

Adding Depth to Microplastics.

The effects and risks of microplastics correlate with three-dimensional (3D) properties, such as the volume and surface area of the biologically accessible fraction of the diverse particle mixtures as they occur in nature. However, these 3D parameters are difficult to estimate because measurement methods for spectroscopic and visible light image analysis yield data in only two dimensions (2D). The best-existing 2D to 3D conversion models require calibration for each new set of particles, which is labor-intensive. Here we introduce a new model that does not require calibration and compare its performance with existing models, including calibration-based ones. For the evaluation, we developed a new method in which the volumes of environmentally relevant microplastic mixtures are estimated in one go instead of on a cumbersome particle-by-particle basis. With this, the new Barchiesi model can be seen as the most universal. The new model can be implemented in software used for the analysis of infrared spectroscopy and visual light image analysis data and is expected to increase the accuracy of risk assessments based on particle volumes and surface areas as toxicologically relevant metrics.

How Digestive Processes Can Affect the Bioavailability of PCBs Associated with Microplastics: A Modeling Study Supported by Empirical Data.

The transfer kinetics of plastic-associated chemicals during intestinal digestive processes is unknown. Here, we assessed whether digestive processes affect chemical exchange kinetics on microplastics, using an in vitro gut fluid digestive model mimicking the human upper intestinal tract. Chemical exchange kinetics of microplastics were measured for 10 polychlorinated biphenyls (PCBs) as proxies for the broad class of hydrophobic organic chemicals. Following earlier studies, olive oil was used as a proxy for digestible food, under high and low digestive enzyme activities. The micelle-water and oil-water partition coefficients of the 10 PCBs were also determined to evaluate the relative contribution of each gut component to sorb PCBs. A new biphasic and reversible chemical exchange model, which included the digestion process, fitted well to the empirical data. We demonstrate that the digestive processes that break down contaminated food can lead to a substantial increase in chemical concentration in microplastics by a factor of 10-20, thereby reducing the overall chemical bioavailability in the gastrointestinal tract when compared to a scenario without microplastics. Higher enzyme activities result in more chemicals being released by the digested food, thereby resulting in higher chemical concentrations in the microplastics. While the model-calibrated kinetic parameters are specific to the studied scenario, we argue that the mechanism of the reduced bioavailability of chemicals and the modeling tool developed have generic relevance. These digestive processes should be considered when assessing the risks of microplastics to humans and also biomagnification in aquatic food webs.

Microplastic Effect Tests Should Use a Standard Heterogeneous Mixture: Multifarious Impacts among 16 Benthic Invertebrate Species Detected under Ecologically Relevant Test Conditions.

Microplastics require a risk assessment framework that takes their multidimensionality into account while exclusively considering robust data. Therefore, effect tests should use a diverse, environmentally relevant microplastic (ERMP) standard material that adheres to high-quality requirements. In this study, we provide chronic dose-effect relationships and effect thresholds for 16 benthic species exposed to ERMP. The ERMP was created from plastic items collected from natural sources and cryogenically milled to represent the diversity of microplastics. The test design met 20 previously published quality assurance and quality control criteria. Adverse effect thresholds (EC10) were determined at ERMP concentrations of 0.11 ± 0.17% sediment dry weight (Gammarus pulex, growth), 0.49 ± 0.68% sediment dry weight (Lumbriculus variegatus, growth), and 1.90 ± 1.08% sediment dry weight (L. variegatus, reproduction). A positive effect of microplastics, such as decreased mortality, was observed for Cerastoderma edule (EC10 = 0.021 ± 0.027% sediment dry weight) and Sphaerium corneum (EC10 = 7.67 ± 3.41% sediment dry weight), respectively. Several of these laboratory-based single-species effect thresholds for ERMP occurred at concentrations lower than those found in the environment. For other species, no significant effects were detected up to an ERMP dose of 10% dry weight.

Maximizing Realism: Mapping Plastic Particles at the Ocean Surface Using Mixtures of Normal Distributions.

Current methods of characterizing plastic debris use arbitrary, predetermined categorizations and assume that the properties of particles are independent. Here we introduce Gaussian mixture models (GMM), a technique suitable for describing non-normal multivariate distributions, as a method to identify mutually exclusive subsets of floating macroplastic and microplastic particles (latent class analysis) based on statistically defensible categories. Length, width, height and polymer type of 6,942 particles and items from the Atlantic Ocean were measured using infrared spectroscopy and image analysis. GMM revealed six underlying normal distributions based on length and width; two within each of the lines, films, and fragments categories. These classes differed significantly in polymer types. The results further showed that smaller films and fragments had a higher correlation between length and width, indicating that they were about the same size in two dimensions. In contrast, larger films and fragments showed low correlations of height with length and width. This demonstrates that larger particles show greater variability in shape and thus plastic fragmentation is associated with particle rounding. These results offer important opportunities for refinement of risk assessment and for modeling the fragmentation and distribution of plastic in the ocean. They further illustrate that GMM is a useful method to map ocean plastics, with advantages over approaches that use arbitrary categorizations and assume size independence or normal distributions.

Potential Artifacts and Control Experiments in Toxicity Tests of Nanoplastic and Microplastic Particles.

To fully understand the potential ecological and human health risks from nanoplastics and microplastics (NMPs) in the environment, it is critical to make accurate measurements. Similar to past research on the toxicology of engineered nanomaterials, a broad range of measurement artifacts and biases are possible when testing their potential toxicity. For example, antimicrobials and surfactants may be present in commercially available NMP dispersions, and these compounds may account for toxicity observed instead of being caused by exposure to the NMP particles. Therefore, control measurements are needed to assess potential artifacts, and revisions to the protocol may be needed to eliminate or reduce the artifacts. In this paper, we comprehensively review and suggest a next generation of control experiments to identify measurement artifacts and biases that can occur while performing NMP toxicity experiments. This review covers the broad range of potential NMP toxicological experiments, such as in vitro studies with a single cell type or complex 3-D tissue constructs, in vivo mammalian studies, and ecotoxicity experiments testing pelagic, sediment, and soil organisms. Incorporation of these control experiments can reduce the likelihood of false positive and false negative results and more accurately elucidate the potential ecological and human health risks of NMPs.

Lifetime Accumulation of Microplastic in Children and Adults.

Human exposure to microplastic is recognized as a global problem, but the uncertainty, variability, and lifetime accumulation are unresolved. We provide a probabilistic lifetime exposure model for children and adults, which accounts for intake via eight food types and inhalation, intestinal absorption, biliary excretion, and plastic-associated chemical exposure via a physiologically based pharmacokinetic submodel. The model probabilistically simulates microplastic concentrations in the gut, body tissue, and stool, the latter allowing validation against empirical data. Rescaling methods were used to ensure comparability between microplastic abundance data. Microplastic (1-5000 µm) median intake rates are 553 particles/capita/day (184 ng/capita/day) and 883 particles/capita/day (583 ng/capita/day) for children and adults, respectively. This intake can irreversibly accumulate to 8.32 × 103 (90% CI, 7.08 × 102-1.91 × 106) particles/capita or 6.4 (90% CI, 0.1-2.31 × 103) ng/capita for children until age 18, and up to 5.01 × 104 (90% CI, 5.25 × 103-9.33 × 106) particles/capita or 40.7 (90% CI, 0.8-9.85 × 103) ng/capita for adults until age 70 in the body tissue for 1-10 µm particles. Simulated microplastic concentrations in stool agree with empirical data. Chemical absorption from food and ingested microplastic of the nine intake media based on biphasic, reversible, and size-specific sorption kinetics, reveals that the contribution of microplastics to total chemical intake is small. The as-yet-unknown contributions of other food types are discussed in light of future research needs.

Automated µFTIR Imaging Demonstrates Taxon-Specific and Selective Uptake of Microplastic by Freshwater Invertebrates.

Microplastic particles can be deposited to sediments and subsequently ingested by benthic organisms. It is unknown to what extent ingestion of microplastic is taxon-specific or whether taxa can be selective toward certain types of microplastics. Here, we used state-of-the-art automated micro-Fourier-transform infrared (µFTIR) imaging and attenuated total reflectance FTIR spectroscopy to determine small-size (20-500 µm) and large-size (500-5000 µm) microplastic particles in sediments and a range of benthic invertebrate species sampled simultaneously from the Dommel River in the Netherlands. Microplastic number concentrations differed across taxa at the same locations, demonstrating taxon-specific uptake, whereas size distributions were the same across sediments and taxa. At the site with the highest concentration, microplastic occupied up to 4.0% of the gut volume of Asellidae. Particle shape distributions were often not statistically different between sediments and taxa, except for Astacidea at one of the locations where the proportion of particles with a length to width ratio >3 (i.e., fibers) was twice as high in sediments than in Astacidea. Acrylates/polyurethane/varnish was predominately found in sediments, while soft and rubbery polymers ethylene propylene diene monomer and polyethylene-chlorinated were the dominant polymers found in invertebrates. Microplastic polymer composition and thus polymer density differed significantly between invertebrates and their host sediment. Trophic transfer at the base of the food web appears to have a filter function with respect to microplastic particle types and shapes. Together with the very high ingestion rates, this has clear implications for ecological and human health risks, where uptake concerns edible species (e.g., Astacidea).

Global Plastic Pollution Observation System to Aid Policy.

Plastic pollution has become one of the most pressing environmental challenges and has received commensurate widespread attention. Although it is a top priority for policymakers and scientists alike, the knowledge required to guide decisions, implement mitigation actions, and assess their outcomes remains inadequate. We argue that an integrated, global monitoring system for plastic pollution is needed to provide comprehensive, harmonized data for environmental, societal, and economic assessments. The initial focus on marine ecosystems has been expanded here to include atmospheric transport and terrestrial and freshwater ecosystems. An earth-system-level plastic observation system is proposed as a hub for collecting and assessing the scale and impacts of plastic pollution across a wide array of particle sizes and ecosystems including air, land, water, and biota and to monitor progress toward ameliorating this problem. The proposed observation system strives to integrate new information and to identify pollution hotspots (i.e., production facilities, cities, roads, ports, etc.) and expands monitoring from marine environments to encompass all ecosystem types. Eventually, such a system will deliver knowledge to support public policy and corporate contributions to the relevant United Nations (UN) Sustainable Development Goals (SDGs).

Quality Criteria for Microplastic Effect Studies in the Context of Risk Assessment: A Critical Review.

In the literature, there is widespread consensus that methods in plastic research need improvement. Current limitations in quality assurance and harmonization prevent progress in our understanding of the true effects of microplastic in the environment. Following the recent development of quality assessment methods for studies reporting concentrations in biota and water samples, we propose a method to assess the quality of microplastic effect studies. We reviewed 105 microplastic effect studies with aquatic biota, provided a systematic overview of their characteristics, developed 20 quality criteria in four main criteria categories (particle characterization, experimental design, applicability in risk assessment, and ecological relevance), propose a protocol for future effect studies with particles, and, finally, used all the information to define the weight of evidence with respect to demonstrated effect mechanisms. On average, studies scored 44.6% (range 20-77.5%) of the maximum score. No study scored positively on all criteria, reconfirming the urgent need for better quality assurance. Most urgent recommendations for improvement relate to avoiding and verifying background contamination, and to improving the environmental relevance of exposure conditions. The majority of the studies (86.7%) evaluated on particle characteristics properly, nonetheless it should be underlined that by failing to provide characteristics of the particles, an entire experiment can become irreproducible. Studies addressed environmentally realistic polymer types fairly well; however, there was a mismatch between sizes tested and those targeted when analyzing microplastic in environmental samples. In far too many instances, studies suggest and speculate mechanisms that are poorly supported by the design and reporting of data in the study. This represents a problem for decision-makers and needs to be minimized in future research. In their papers, authors frame 10 effects mechanisms as "suggested", whereas 7 of them are framed as "demonstrated". When accounting for the quality of the studies according to our assessment, three of these mechanisms remained. These are inhibition of food assimilation and/or decreased nutritional value of food, internal physical damage, and external physical damage. We recommend that risk assessment addresses these mechanisms with higher priority.

Solving the Nonalignment of Methods and Approaches Used in Microplastic Research to Consistently Characterize Risk.

The lack of standard approaches in microplastic research limits progress in the abatement of plastic pollution. Here, we propose and test rescaling methods that are able to improve the alignment of methods used in microplastic research. We describe a method to correct for the differences in size ranges as used by studies reporting microplastic concentrations and demonstrate how this reduces the variation in aqueous-phase concentrations caused by method differences. We provide a method to interchange between number, volume, and mass concentrations using probability density functions that represent environmental microplastic. Finally, we use this method to correct for the incompatibility of data as used in current species sensitivity distributions (SSDs), caused by differences in the microplastic types used in effect studies and those in nature. We derived threshold effect concentrations from such a corrected SSD for freshwater species. Comparison of the rescaled exposure concentrations and threshold effect concentrations reveals that the latter would be exceeded for 1.5% of the known surface water exposure concentrations worldwide. Altogether, this toolset allows us to correct for the diversity of microplastic, to address it in a common language, and to assess its risks as one environmental material.

Modeling Decreased Resilience of Shallow Lake Ecosystems toward Eutrophication due to Microplastic Ingestion across the Food Web.

The discovery of microplastic (MP) being present in freshwaters has stimulated research on the impacts of MP on freshwater organisms. To date, research has focused on primary effects, leaving questions with respect to secondary effects at the level of freshwater food webs unanswered. Here, we use a theoretical modeling approach to investigate the hypothesis that MP imposes negative impacts on the level of freshwater shallow lake food webs. We find that increasing MP levels have the potential to affect the critical phosphorus loading (CPL), which is defined as the threshold for regime shifts between clear and turbid states of the water column. The possible occurrence of catastrophic cascades due to MP pollution is predominantly driven by the negative effects of MP on zooplankton. We explore the possible states of the food web by scenario analysis and show that the secondary effects of MP at current concentrations are likely to be negligible. However, at the current rate of MP production, a 20-40% reduction in the CPL would occur by the end of this century, suggesting a loss of resilience in shallow lakes that would be subject to abrupt changes in the food web under lower nutrient loading.

Transfer of PCBs from Microplastics under Simulated Gut Fluid Conditions Is Biphasic and Reversible.

The role of plastic as a vector for bioaccumulation of toxic chemicals is central to the risk assessment of microplastic for human health and the environment. However, transfer kinetics of sorbed contaminants from ingested microplastics are poorly understood. We develop and parametrize a chemical exchange model on microplastics in a gut fluid mimic of aquatic biota, and also included food to provide a better representation of contaminant dynamics when plastic and food are ingested, as would occur in nature. The transfer kinetics of 14 polychlorinated biphenyls (PCBs) were measured in gut fluid mimic systems under three environmentally relevant exposure scenarios of plastic ingestion by organisms, for low-density polyethylene (LDPE) and polyvinyl chloride (PVC), and were evaluated with the model. Chemical transfer was demonstrated to be biphasic and fully reversible, with fast exchange within hours followed by a slow transfer lasting for weeks to months. In clean gut systems, the bioavailability of plastic-associated PCBs for lugworms and cod ranged from 14 to 42% and 45-83% respectively. However, in contaminated gut systems, clean microplastic was capable of rapidly extracting ("cleaning") PCBs from food inside the gut, thus demonstrating that the effect of microplastic is context dependent. Therefore, chemical contamination and cleaning are likely to occur simultaneously due to the ingestion of microplastic.

Accumulation of Plastic Debris and Associated Contaminants in Aquatic Food Webs.

We present a generic theoretical model (MICROWEB) that simulates the transfer of microplastics and hydrophobic organic chemicals (HOC) in food webs. We implemented the model for an Arctic case comprised of nine species including Atlantic cod and polar bear as top predator. We used the model to examine the effect of plastic ingestion on trophic transfer of microplastics and persistent HOCs (PCBs) and metabolizable HOCs (PAHs), spanning a wide range of hydrophobicities. In a scenario where HOCs in plastic and water are in equilibrium, PCBs biomagnify less when more microplastic is ingested, because PCBs biomagnify less well from ingested plastic than from regular food. In contrast, PAHs biomagnify more when more microplastic is ingested, because plastic reduces the fraction of PAHs available for metabolization. We also explore nonequilibrium scenarios representative of additives that are leaching out, as well as sorbing HOCs, quantitatively showing how the above trends are strengthened and weakened, respectively. The observed patterns were not very sensitive to modifications in the structure of the food web. The model can be used as a tool to assess prospective risks of exposure to microplastics and complex HOC mixtures for any food web, including those with relevance for human health.

Quality Criteria for the Analysis of Microplastic in Biota Samples: A Critical Review.

Data on ingestion of microplastics by marine biota are quintessential for monitoring and risk assessment of microplastics in the environment. Current studies, however, portray a wide spread in results on the occurrence of microplastic ingestion, highlighting a lack of comparability of results, which might be attributed to a lack of standardization of methods. We critically review and evaluate recent microplastic ingestion studies in aquatic biota, propose a quality assessment method for such studies, and apply the assessment method to the reviewed studies. The quality assessment method uses ten criteria: sampling method and strategy, sample size, sample processing and storage, laboratory preparation, clean air conditions, negative controls, positive controls, target component, sample (pre)treatment, and polymer identification. The results of this quality assessment show a dire need for stricter quality assurance in microplastic ingestion studies. On average, studies score 8.0 out of 20 points for "completeness of information" and 0 for "reliability". Alongside the assessment method, a standardized protocol for detecting microplastic in biota samples incorporating these criteria is provided.

Microplastic Effect Thresholds for Freshwater Benthic Macroinvertebrates.

Now that microplastics have been detected in lakes, rivers, and estuaries all over the globe, evaluating their effects on biota has become an urgent research priority. This is the first study that aims at determining the effect thresholds for a battery of six freshwater benthic macroinvertebrates with different species traits, using a wide range of microplastic concentrations. Standardized 28 days single species bioassays were performed under environmentally relevant exposure conditions using polystyrene microplastics (20-500 µm) mixed with sediment at concentrations ranging from 0 to 40% sediment dry weight (dw). Microplastics caused no effects on the survival of Gammarus pulex, Hyalella azteca, Asellus aquaticus, Sphaerium corneum, and Tubifex spp. and no effects were found on the reproduction of Lumbriculus variegatus. No significant differences in growth were found for H. azteca, A. aquaticus, S. corneum, L. variegatus, and Tubifex spp. However, G. pulex showed a significant reduction in growth (EC10 = 1.07% sediment dw) and microplastic uptake was proportional with microplastic concentrations in sediment. These results indicate that although the risks of environmentally realistic concentrations of microplastics may be low, they still may affect the biodiversity and the functioning of aquatic communities which after all also depend on the sensitive species.

Ingestion and Chronic Effects of Car Tire Tread Particles on Freshwater Benthic Macroinvertebrates.

Micronized particles released from car tires have been found to contribute substantially to microplastic pollution, triggering the need to evaluate their effects on biota. In the present study, four freshwater benthic macroinvertebrates were exposed for 28 days to tread particles (TP; 10-586 µm) made from used car tires at concentrations of 0, 0.1, 0.3, 1, 3, and 10% sediment dry weight. No adverse effects were found on the survival, growth, and feeding rate of Gammarus pulex and Asellus aquaticus, the survival and growth of Tubifex spp., and the number of worms and growth of Lumbriculus variegatus. A method to quantify TP numbers inside biota was developed and here applied to G. pulex. In bodies and faces of G. pulex exposed to 10% car tire TP, averages of 2.5 and 4 tread particles per organism were found, respectively. Chemical analysis showed that, although car tire TP had a high intrinsic zinc content, only small fractions of the heavy metals present were bioavailable. PAHs in the TP-sediment mixtures also remained below existing toxicity thresholds. This combination of results suggests that real in situ effects of TP and TP-associated contaminants when dispersed in sediments are probably lower than those reported after forced leaching of contaminants from car tire particles.

Pollutants in Plastics within the North Pacific Subtropical Gyre.

Here we report concentrations of pollutants in floating plastics from the North Pacific accumulation zone (NPAC). We compared chemical concentrations in plastics of different types and sizes, assessed ocean plastic potential risks using sediment quality criteria, and discussed the implications of our findings for bioaccumulation. Our results suggest that at least a fraction of the NPAC plastics is not in equilibrium with the surrounding seawater. For instance, "hard plastic" samples had significantly higher PBDE concentrations than "nets and ropes" samples, and 29% of them had PBDE composition similar to a widely used flame-retardant mixture. Our findings indicate that NPAC plastics may pose a chemical risk to organisms as 84% of the samples had at least one chemical exceeding sediment threshold effect levels. Furthermore, our surface trawls collected more plastic than biomass (180 times on average), indicating that some NPAC organisms feeding upon floating particles may have plastic as a major component of their diets. If gradients for pollutant transfer from NPAC plastic to predators exist (as indicated by our fugacity ratio calculations), plastics may play a role in transferring chemicals to certain marine organisms.

Ups and Downs in the Ocean: Effects of Biofouling on Vertical Transport of Microplastics.

Recent studies suggest size-selective removal of small plastic particles from the ocean surface, an observation that remains unexplained. We studied one of the hypotheses regarding this size-selective removal: the formation of a biofilm on the microplastics (biofouling). We developed the first theoretical model that is capable of simulating the effect of biofouling on the fate of microplastic. The model is based on settling, biofilm growth, and ocean depth profiles for light, water density, temperature, salinity, and viscosity. Using realistic parameters, the model simulates the vertical transport of small microplastic particles over time, and predicts that the particles either float, sink to the ocean floor, or oscillate vertically, depending on the size and density of the particle. The predicted size-dependent vertical movement of microplastic particles results in a maximum concentration at intermediate depths. Consequently, relatively low abundances of small particles are predicted at the ocean surface, while at the same time these small particles may never reach the ocean floor. Our results hint at the fate of "lost" plastic in the ocean, and provide a start for predicting risks of exposure to microplastics for potentially vulnerable species living at these depths.

The Effect of Microplastic on the Uptake of Chemicals by the Lugworm Arenicola marina (L.) under Environmentally Relevant Exposure Conditions.

It has been hypothesized that ingestion of microplastic increases exposure of aquatic organisms to hydrophobic contaminants. To date, most laboratory studies investigated chemical transfer from ingested microplastic without taking other exposure pathways into account. Therefore, we studied the effect of polyethylene (PE) microplastic in sediment on PCB uptake by Arenicola marina as a model species, quantifying uptake fluxes from all natural exposure pathways. PCB concentrations in sediment, biota lipids (Clip) and porewater measured with passive samplers were used to derive lipid-normalized bioaccumulation metrics Clip, Biota sediment accumulation factor (BSAF), Bioaccumulation factor (BAF) and the Biota plastic accumulation factor (BPAF). Small effects of PE addition were detected suggesting slightly increased or decreased bioaccumulation. However, the differences decreased in magnitude dependent on the metric used to assess bioaccumulation, in the order: Clip > BSAF > BPAF > BAF, and were nonsignificant for BAF. The fact that BAF, that is, normalization of Clip on porewater concentration, largely removed all effects of PE, shows that PE did not act as a measurable vector of PCBs. Biodynamic model analysis confirmed that PE ingestion contributed marginally to bioaccumulation. This work confirmed model-based predictions on the limited relevance of microplastic for bioaccumulation under environmentally realistic conditions, and illustrated the importance of assessing exposure through all media in microplastic bioaccumulation studies.