Ecotoxicity Tests Research Articles

The effects of biosynthesized silver nanoparticle on oxidative stress parameters in model organism Artemia salina (Linnaeus, 1758)

Evaluation of the effects of nanomaterials in various habitats and living groups is increasing with in vitro and in vivo studies. This study aims to describe a biological procedure to synthesize silver nanoparticles using an extract of Marrubium astracanicum and to determine the aquatic environment’s oxidative stress on Artemia salina. Silver nanoparticle formation was visually observed with color change. Its structural properties, such as size, shape, morphology, and stability, were characterized by UV-Vis spectrophotometer, TEM, SEM, XRD, and DLS analyses. TEM image showed that the synthesized AgNPs have a size of between 20 and 50 nm. The effects of biosynthesized silver nanoparticles at different concentrations (control,0.2, 1, 5, 10, 25, and 50 mg/mL) and exposure times (24, 48 and 72 h) on total glutathione (GSH) and lipid peroxidation levels (MDA) were investigated on the Artemia salina organism, which is one of the most common saltwater organisms frequently used in ecotoxicity tests.

Comprehensive Assessment of Lead Bioaccumulation in Helix aspersa (Müller, 1774), Snails: A Study of Histopathological and Biochemical Impacts

Background: The snail Helixa spersa is considered a relevant bioindicator of soil pollution by metallic elements due to its resistance and accumulation capabilities. Methods: This study aims to determine the dose-response relationship between different concentrations of lead and its toxic effects on juvenile H. aspersa through a semi-static ecotoxicity test under controlled conditions. In addition, continuous monitoring of carbohydrates, lipids and proteins of the snails affected by this metal was evaluated along with a histopathological study of the hepatopancreas. Result: The two-month lead exposure in H. aspersa resulted in significant alterations in biochemical and histological parameters. Lead concentrations in the hepatopancreas and foot exhibited a dose-dependent increase, with higher levels observed in the former. Elevated lead concentrations (1000 ìg/g and 1500 ìg/g of soil) led to substantial protein increase in both organs, while even lower doses displayed considerable protein elevation in the hepatopancreas. Carbohydrate levels were significantly lower in all contaminated groups compared to controls. Increasing lead concentrations caused a reduction in lipid levels. Histological analysis revealed distinct alterations, including excretory cell hypertrophy, tubule clustering and inflammatory changes in the hepatopancreas of treated snails. Severe histological damage, including inflammatory infiltrates, cellular debris and necrosis, were observed at higher lead concentrations (1000 and 1500 ìg/g). These findings emphasize the impact of lead exposure on biochemical profiles and organ histology in H. aspersa, highlighting the potential ecological implications of heavy metal contamination.

Testing the Aquatic Toxicity of 2D Few-Layer Graphene Inks Using Rainbow Trout (Oncorhynchus mykiss): In Vivo and In Vitro Approaches to Support an SSbD Assessment.

Graphene-based conductive inks offer attractive possibilities in many printing technology applications. Often, these inks contain a mixture of compounds, such as solvents and stabilizers. For the safe(r) and sustainable use of such materials in products, potentially hazardous components must be identified and considered in the design stage. In this study, the hazards of few-layer graphene (FLG)-based ink formulations were tested in fish using in vitro (RTL-W1 cell line) and in vivo aquatic ecotoxicity tests (OECD TG 203). Five ink formulations were produced using different processing steps, containing varying amounts of solvents and stabilizers, with the end products formulated either in aqueous solutions or in powder form. The FLG ink formulations with the highest contents of the stabilizer sodium deoxycholate showed greater in vitro cytotoxic effects, but they did not provoke mortality in juvenile rainbow trout. However, exposure led to increased activities of the cytochrome P450 1a (Cyp1a) and Cyp3a enzymes in the liver, which play an essential role in the detoxification of xenobiotics, suggesting that any effects will be enhanced by the presence of the stabilizers. These results highlight the importance of an SSbD approach together with the use of appropriate testing tools and strategies. By incorporating additional processing steps to remove identified cytotoxic residual solvents and stabilizers, the hazard profile of the FLG inks improved, demonstrating that, by following the principles of the European Commission's safe(r) and sustainable by design (SSbD) framework, one can contribute to the safe(r) and sustainable use of functional and advanced 2D materials in products.

Ecotoxicological Effects of Potassium Dichromate on the Tadpole Shrimp Triops longicaudatus

The tadpole shrimp Triops longicaudatus is a freshwater crustacean with fast embryonic and larval development, short life cycle, and high fecundity. They are very active swimmers of a reasonable size, easy to spot and record. Such characteristics make it a promising candidate as an experimental model in ecotoxicology to evaluate the effects of aquatic pollutants, particularly using its locomotor behavior as an endpoint. To evaluate the sensitivity of T. longicaudatus and develop endpoints of interest, we conducted exposure experiments with lethal and sub-lethal concentrations of potassium dichromate, a compound known for its ecotoxicological importance and as a hexavalent chromium source. The endpoints evaluated were mortality, growth, sexual maturation, reproductive output, cholinesterase activity and locomotor/swimming behavior. The 96 h median lethal concentration was found to be 65 µg/L. Furthermore, exposure to potassium dichromate at higher concentrations had a significant negative impact on the growth rate of T. longicaudatus in terms of both body mass and length. The time for maturation was also delayed at higher concentrations. In addition, locomotor behavior allowed for the discrimination of all tested chromium concentrations and the control group and from each other, proving to be the most sensitive endpoint. Overall, the data support the potential of T. longicaudatus as a model for ecotoxicity testing, using apical endpoints with impact at the population level; in particular, results suggest that behavior assessments in this species might be useful for detecting hazardous compounds in environmental monitoring of freshwater ecosystems.

Natural products as a control measure of the Achatina fulica (Gastropoda: Achatinidae).

Achatina fulica is a terrestrial mollusk known as the giant African snail that is related to environmental, economic, urban, and public health problems. As control measures for this mollusk, cooking salt (NaCl) and calcium oxide (CaO) are used, and baits are composed of metaldehyde. However, these measures have environmental toxicity and impact the soil. In this way, natural products have been tested on this mollusk to discover and develop a substance to combat this urban and agricultural pest. This article aims to evaluate studies involving natural products to control the population of Achatina fulica. Articles and works published in books were included in the present work. A total of 1,103 works were found during the search. Of these, 14 works met the objective of these review and were included in this article. The tests do not possess methodological standardization, do not have a maximum concentration to be considered active, or a maximum exposure time. A lack of standardization in the methodology of tests on A. fulica was observed. The performance of tests on other life stages of the mollusk, as well as tests that analyze other parameters, are essential. Only one article analyzed presented phytochemical analysis. No ecotoxicity tests were reported either. Some extracts showed promising results, highlighting the aqueous extract of Capsicum frutescens. More studies investigating the molluscicidal activity of natural products on A. fulica are needed. It is very relevant that the new studies present a phytochemical analysis of the tested extracts, as well as ecotoxicity studies.

Ecotoxicity testing of nanomaterials in sediment – suggestions to improve science and regulation

Sediments generally act as a sink of released manufactured nanomaterials (NMs).

Proposing Effective Ecotoxicity Test Species for Chemical Safety Assessment in East Asia: A Review.

East Asia leads the global chemical industry, but environmental chemical risk in these countries is an emerging concern. Despite this, only a few native species that are representative of East Asian environments are listed as test species in international guidelines compared with those native to Europe and America. This review suggests that Zacco platypus, Misgurnus anguillicaudatus, Hydrilla verticillata, Neocaridina denticulata spp., and Scenedesmus obliquus, all resident to East Asia, are promising test species for ecotoxicity tests. The utility of these five species in environmental risk assessment (ERA) varies depending on their individual traits and the state of ecotoxicity research, indicating a need for different applications of each species according to ERA objectives. Furthermore, the traits of these five species can complement each other when assessing chemical effects under diverse exposure scenarios, suggesting they can form a versatile battery for ERA. This review also analyzes recent trends in ecotoxicity studies and proposes emerging research issues, such as the application of alternative test methods, comparative studies using model species, the identification of specific markers for test species, and performance of toxicity tests under environmentally relevant conditions. The information provided on the utility of the five species and alternative issues in toxicity tests could assist in selecting test species suited to study objectives for more effective ERA.

Effluent toxicity study using biomarkers for ciprofloxacin photoelectrocatalytic degradation by bismuth-doped titanium dioxide nanotubes

ABSTRACT Ciprofloxacin hydrochloride (CIP) is a broad-spectrum synthetic antibiotic often found in domestic sewage and industrial waste due to the inefficiency of conventional treatments. Given the potential risk of drug accumulation, this study presents coatings of titanium dioxide nanotubes (TiO2) doped with different bismuth (Bi) concentrations to degrade CIP through photocatalytic and photoelectrochemical processes. Characterization studies revealed that bismuth (Bi) doping affected the morphology of the materials, with concentrations of 0.01 and 0.05 mol L−1, resulting in collapsed materials with a smaller active surface area. Photocatalysis tests for all the materials exhibited a similar degree of efficiency to photolysis, approximately 33%. Ecotoxicity tests using the biomarkers Lactuca sativa L., Lemna minor, and Artemia salina indicated that, although they were similar to photolysis in terms of efficiency, the effluents generated when employing the doped catalysts showed lower levels of toxicity, with the best results achieved for the material doped with 0.005 mol L−1 of Bi, with a toxicity level approximately 40% lower. Photoelectrocatalysis proved to be the most efficient CIP degradation technique. The highest degradation rate was observed for materials doped with 0.005 mol L−1 of Bi, with an efficiency of 46%, which is 1.4 times more efficient than photolysis. These results demonstrate that materials doped with low amounts of Bi can be effectively used as photoanodes for drug degradation, as their performance is superior, and the final product generated exhibits low toxicity to living organisms.

Removal of Rhodamine B dye by adsorption onto an eco-friendly zeolite and machine learning modeling

The present work aims to synthesize and characterize a zeolite doped with cobalt (ZO-Co) from such as rice husk, residual sludge and pumice stone for application in the removal of RhB dye by adsorption. Experimental design and machine learning were used to verify the ideal condition of RhB adsorption, as well, as ecotoxicity tests against Lactuca sativa L. seeds were carried out. ZO-Co showed diffraction peaks of the tsaregorodsevite phase, groups containing hydroxyl, silicates and aluminosilicates. The ideal condition by machine learning (ML) was [RhB] of = 130 mg/L, [ZO-Co] = 0.6 g/L, pH ≈ 7, and T = 298.15±2 K. The best experimental fit for the adsorption data was the isotherm of Khan (equilibrium) and Avrami (kinetic model), where the maximum adsorption capacity was 259.17 mg g−1 with 83.66 % of the RhB removal. It is noteworthy that in ecotoxicity tests ZO-Co showed root radicular growth of 29.74 mm in 7 days. Therefore, cobalt-doped ZO can be effectively used for the adsorption by organic pollutant RhB correlating topics of environment and nanotechnology.

Polymers of low concern? Assessment of microplastic particles used in 3D printing regarding their toxicity on Raphidocelis subcapitata and Daphnia magna

The potential effects of microplastic particle exposure on environmental organisms has sparked intense research activities. Various studies have been conducted, however on a limited set of mostly pristine polymer materials. In parallel to the ongoing research activities, it is discussed to include non-natural polymers into the registration process under REACH. Currently, non-natural polymers are exempted from registration, based on the general assumption of being non-hazardous due to their high molecular weight. In addition, the extensive number of polymers exceeding the registration capacities was mentioned as a reason for exemption. Hence, relevant polymers requiring a registration shall be selected according to specific criteria that help to identify those with a concern for hazardous effects. In the line of these developments we here present the results of a systematic ecotoxicity testing of 16 microplastic particles of different polymer composition, part of which are micronized polymer powders used in 3D printing (intentionally produced primary microplastic, losing particle shape by the 3D printing). All polymer materials were systematically varied and extensively characterised with regard to their properties (e.g. particle size, cross-linking, molar mass distribution, end groups, reactivity). Most of the polymers exerted toxicity in 48 h acute Daphnia magna immobilisation assay and 72 h chronic Raphidocelis subcapitata growth inhibition assay, except one PA and one HDPE material showing toxicity in D. magna, and one HDPE material showing toxicity in green algae. From these results we conclude that none of the microplastic particles studied here, independent of their polymer properties, give raise to concern for hazardous effects.

Defining the data gap: What do we know about environmental exposure, hazards and risks of pharmaceuticals in the European aquatic environment?

Active pharmaceutical ingredients (APIs) and their transformation products inevitably enter waterways where they might cause adverse effects to aquatic organisms. Identifying the potential risks of APIs in the environment is therefore a goal and current strategic direction of environmental management described in the EU Strategic Approach to Pharmaceuticals in the Environment and the Green Deal. This is challenged by a paucity of monitoring and ecotoxicity data to adequately describe risks.In this study we analyze measured environmental concentrations (MECs) of APIs from 5933 sites in 25 European countries as documented in the EMPODAT database or collected by the German Environment Agency for the time period between 1997 and 2020. These data were compared with empirical data on the ecotoxicity of APIs from the U.S. EPA ECOTOX database. Although 1763 uniquely identifiable APIs are registered with the European Medicines Agency (EMA) for sale in the European Economic Area (EEA), only 312 (17.7%) of these are included in publicly available monitoring data, 36 (1.8%) compounds have sufficient ecotoxicological data to derive a PNEC, and only 27 (1.5%) compounds meet both the hazard and exposure data requirements required to to perform an environmental risk assessment according to EMA guidelines. Four of these compounds (14.8%) had a median risk quotient (RQ) > 1. Endocrine disruptors had the highest median RQ, with 7.0 and 5.6 for 17α-ethinyl-estradiol and 17β-estradiol respectively.A comparison of in-silico and empirical exposure data for 72 APIs demonstrated the high protectiveness of the current EMA guidelines, with predicted environmental concentrations (PECs) exceeding median MECs in 98.6% of cases, with a 100-fold median increase.This study describes the data shortfalls hindering an accurate assessment of the risk posed to European waterways by APIs, and identifies 68 APIs for prioritized inclusion in monitoring programs, and 66 APIs requiring ecotoxicity testing to fill current data gaps.

Comprehensive assessment of health and ecological risk of cadmium in agricultural soils across China: A tiered framework

Soil cadmium (Cd) contamination has been increasingly serious in agricultural land across China, posing unexpected risks to human health concerning crop safety and terrestrial ecosystems. This study collected Cd concentration data from 3388 soil sites in agricultural regions. To assess the Cd risk to crop safety, a comprehensive sampling investigation was performed to develop reliable Soil Plant Transfer (SPT) model. Eco-toxicity tests with representative soils and organism was conducted to construct the Species Sensitivity Distribution (SSD) for ecological risk assessment. Then, a tiered framework was applied based on Accumulation index, deterministic method (Hazard quotient), and probabilistic assessment (Monte Carlo and Joint Probability Curve). The results revealed the widespread Cd enrichment in agricultural soils, mainly concentrated in Central, Southern, and Southwest China. Risk assessments demonstrated the greater risks related to crop safety, while the ecological risks posed by soil Cd were manageable. Notably, agricultural soils in southern regions of China exhibited more severe risks to both crop safety and soil ecosystem, compared to other agricultural regions. Furthermore, tiered methodology proposed here, can be adapted to other trace elements with potential risks to crop safety and terrestrial ecosystem.

Sustainable and biodegradable poly(butylene succinate‐co‐terephthalate)/poly(propylene carbonate) blown films with enhanced compatibility, mechanical, and barrier properties

AbstractIn this study, a novel environment‐friendly PBST/PPC‐based blown film was prepared using maleic anhydride (MA) as a reactive compatibilizer to enhance the compatibility between poly(butylene succinate‐co‐terephthalate) (PBST) and poly(propylene carbonate) (PPC). Results of rheological testing and gel permeation chromatography (GPC) indicated that MA reacted with PBST/PPC during melt‐blending extrusion. Morphological analysis of the cryo‐fractured surfaces of PBST/PPC blend showed significantly improved compatibility between PBST and PPC with the addition of MA. Moreover, the Young's modulus, tensile strength, breaking strain, and tear strength of PBST/PPC/MA blown films increased with an increase in MA content. In comparison to PBST/MA blown film without PPC, the barrier property of PBST/PPC/MA blown films was improved. In addition, in vitro cell experiments showed that the PBST/PPC/MA blown film was suitable for the growth of mouse fibroblast (L929) cells. In vitro ecotoxicity testing on mung bean plant showed that the extracts from the PBST/PPC/MA blown film had no negative effects on the development of mung bean plant. Furthermore, degradability testing in soil also proved that the PBST/PPC/MA blown film had good biodegradability. Thus, the PBST/PPC/MA blown film can be used in fields, such as food packaging and agricultural mulch film.

Nano-Management Approaches for Salt Tolerance in Plants under Field and In Vitro Conditions

Soil salinity is a serious global problem that threatens a high percentage of the global soils. Salinity stress can create ionic, oxidative, and osmotic stress, along with hormonal imbalances, in stressful plants. This kind of stress was investigated on agricultural productivity at different levels, starting in vitro (plant tissue culture), through hydroponics, pots, and field conditions. Several approaches were studied for managing salinity stress, including using traditional materials (e.g., gypsum, sulfur), organic amendments (e.g., compost, biochar, chitosan), and applied manufactured or engineered nanomaterials (NMs). Application of nanomaterials for ameliorating salinity stress has gained great attention due to their high efficiency, eco-friendliness, and non-toxicity, especially biological nanomaterials. The application of NMs did not only support growing stressful plants under salinity stress but also increased the yield of crops, provided an economically feasible nutrient management approach, and was environmentally robust for sustainable crop productivity. Nano-management of salinity may involve applying traditional nano-amendments, biological nanomaterials, nano-enabled nutrients, nano-organic amendments, derived smart nanostructures, and nano-tolerant plant cultivars. Producing different plant cultivars that are tolerant to salinity can be achieved using conventional breeding and plantomics technologies. In addition to the large-scale use of nanomaterials, there is an urgent need to address and treat nanotoxicity. This study aims to contribute to this growing area of research by exploring different approaches for nano-management of current practices under salinity stress under field and in vitro conditions. This study also raises many questions regarding the expected interaction between the toxic effects of salinity and NMs under such conditions. This includes whether this interaction acts positively or negatively on the cultivated plants and soil biological activity, or what regulatory ecotoxicity tests and protocols should be used in research.

Ecotoxicity characterization assisted performance assessment of electro-bioremediation reactors for nitrate and arsenite elimination.

The performance of combined reduction of nitrate (NO3 - ) to dinitrogen gas (N2 ) and oxidation of arsenite (As[III]) to arsenate (As[V]) by a bioelectrochemical system was assessed, supported by ecotoxicity characterization. For the comprehensive toxicity characterization of the untreated model groundwater and the treated reactor effluents, a problem-specific ecotoxicity test battery was established. The performance of the applied technology in terms of toxicity and target pollutant elimination was compared and analyzed. The highest toxicity attenuation was achieved under continuous flow mode with hydraulic retention time (HRT) = 7.5 h, with 95%, nitrate removal rate and complete oxidation of arsenite to arsenate. Daphnia magna proved to be the most sensitive test organism. The results of the D. magna lethality test supported the choice of the ideal operational conditions based on chemical data analysis. The outcomes of the study demonstrated that the applied technology was able to improve the groundwater quality in terms of both chemical and ecotoxicological characteristics. The importance of ecotoxicity evaluation was also highlighted, given that significant target contaminant elimination did not necessarily lower the environmental impact of the initial, untreated medium, in addition, anomalies might occur during the technology operational process which in some instances, could result in elevated toxicity levels.

Microplastics: toxicity and Bioacumulation at the first trophic levels of the food chain

Microplastics are dispersed in nature in all planetary compartments, and can be found in the most remote places of the Ecosystem. They are currently indicated as markers of the Anthropocene, a geological era caused by human changes on the globe. Even though it is found in all environmental compartments, there are still gaps in scientific knowledge about its real impact on living cells and their dynamics, including in the human organism. Therefore, the objective of the study is to use an internationally standardized and standardized method in representative organisms of the ecosystem, thus using the acute and chronic ecotoxicity test with the microcrustacean Daphnia magna and for the algae Desmodesmus subspicatus, respectively, against polyethylene microspheres, a thermoplastic widely used as an exfoliation product and applied in various cosmetics and hygiene materials. They were tested in neonates according to the reference standards, but 30-day-old adult organisms were also exposed. The results obtained, for the crustaceans, show the Effective Concentration that causes effect in 50% of the organisms (EC50%), with 95% confidence of 0.07 g/L and a DSC – Observed effect concentration of 0.025 g/L for neonates. For the adult specimens, 0.025 g/L of EC50 and 0.0025 g/L of DSC were found, showing that the pups are 10x more resistant to plasticizers compared to the adults. For the algae test, the results showed a DSC of 0.05 g/L, inhibiting 20.12% of algae growth compared to the control. In the optical microscope of the crustaceans after exposure, it showed bioaccumulation of the polyethylene microsphere in the carapace and digestive tract of the bioindicator, showing that the impact of microplastics begins in the first trophic levels of the food chain.

ECOTOXICOLOGICAL TESTS OF THE PARTICLEBOARDS CONTAINING RUBBER WASTE

The article is focused on the production and environmental evaluation of wood composites using waste rubber in the construction industry. Used aqueous extracts were prepared from theexperimental wooden composites with various additions of thewaste rubber from tires and waste seals. The pH value and organic pollution (by COD) were determined in the aqueous extracts. The effect on the environmental components (aquatic and terrestrial) was ecotoxicologically tested using the test organisms Sinapis alba, Lemna minorand Daphnia magna. Preliminary acute ecotoxicity tests were performed.

Association between heavy metal uptake and growth and reproduction in the anecic earthworm, Alma nilotica (Grube 1855).

Elevated heavy metal concentrations in soils are a cause for concern as they are hazardous to soil organisms including earthworms which are considered as ecosystem engineers. Current ecotoxicity tests predominantly use temperate earthworm species, and thus there is the need to include a broader genera of native species to improve ecological risk assessment. Alma nilotica, is a tropical anecic earthworm species that survives well under laboratory conditions and has potential for use in ecotoxicology testing but lacks published toxicity data for important pollutants. Growth and reproduction bioassays were carried out with A. nilotica to determine the relationship between the concentrations of Cu, Zn, Pb and Cr in spiked soils and their bioaccumulation and toxic effects. Positive linear relationships were found between soil-metal and internal earthworm-metal concentrations. Cu did not inhibit growth up to 35 days of exposure but became toxic with longer exposure duration. Zn was not regulated by A. nilotica although it is an essential metal that is well regulated by Eisenia sp. commonly used in standard ecotoxicity tests, showing differences in metal regulation by earthworms of different ecological categories. Based on bioaccumulation factors (BAFs), growth inhibition and reproduction effects the metals were ranked in decreasing toxicity as Pb > Cr > Zn > Cu. The mean 20% Internal Effects Concentrations (IEC20s) for reproduction were 1.04, 2.9, 8.3 and 224.2 mg metal kg-1 earthworm for Pb, Cr, Zn and Cu respectively. These data can contribute to the improvement of metal risk assessment particularly in tropical contexts.

Advancements, Challenges, and Future Directions in Aquatic Life Criteria Research in China.

Aquatic life criteria (ALC) serve as the scientific foundation for establishing water quality standards, and in China, significant strides have been made in the development of freshwater ALC. This comprehensive review traces the evolution of China's WQC, focusing on the methodological advancements and challenges in priority pollutants selection, test organism screening, and standardized ecotoxicity testing protocols. It also provides a critical evaluation of quality assurance measures, data validation techniques, and minimum data requirements essential for ALC assessments. The paper highlights China's technical guidelines for deriving ALC, and reviews the published values for typical pollutants, assessing their impact on environmental quality standards. Emerging trends and future research avenues are discussed, including the incorporation of molecular toxicology data and the development of predictive models for pollutant toxicity. The review concludes by advocating for a tiered WQC system that accommodates China's diverse ecological regions, thereby offering a robust scientific basis for enhanced water quality management.

Exposure protocol for ecotoxicity testing of microplastics and nanoplastics.

Despite the increasing concern about the harmful effects of micro- and nanoplastics (MNPs), there are no harmonized guidelines or protocols yet available for MNP ecotoxicity testing. Current ecotoxicity studies often use commercial spherical particles as models for MNPs, but in nature, MNPs occur in variable shapes, sizes and chemical compositions. Moreover, protocols developed for chemicals that dissolve or form stable dispersions are currently used for assessing the ecotoxicity of MNPs. Plastic particles, however, do not dissolve and also show dynamic behavior in the exposure medium, depending on, for example, MNP physicochemical properties and the medium's conditions such as pH and ionic strength. Here we describe an exposure protocol that considers the particle-specific properties of MNPs and their dynamic behavior in exposure systems. Procedure 1 describes the top-down production of more realistic MNPs as representative of MNPs in nature and particle characterization (e.g., using thermal extraction desorption-gas chromatography/mass spectrometry). Then, we describe exposure system development for short- and long-term toxicity tests for soil (Procedure 2) and aquatic (Procedure 3) organisms. Procedures 2 and 3 explain how to modify existing ecotoxicity guidelines for chemicals to target testing MNPs in selected exposure systems. We show some examples that were used to develop the protocol to test, for example, MNP toxicity in marine rotifers, freshwater mussels, daphnids and earthworms. The present protocol takes between 24 h and 2 months, depending on the test of interest and can be applied by students, academics, environmental risk assessors and industries.