An Integrated Testing Strategy for Ecotoxicity (ITS-ECO) Assessment in the Marine Environmental Compartment using Mytilus spp.: A Case Study using Pristine and Coated CuO and TiO2 Nanomaterials.

An integrated testing strategy for ecotoxicity assessment (ITS-ECO) was developed to aid in the hazard and fate assessment of engineered nanomaterials (ENMs) deposited in marine environments using the bivalve Mytilus spp. as a test species. The ENMs copper(II) oxide (CuO) and titanium dioxide (TiO2 ), either in pristine form (core) or with functionalized coatings (polyethylene glycol [PEG], carboxyl [COOH], and ammonia [NH3 ]) were selected as case study materials based on their production levels and use. High-throughput in vitro testing in Tier 1 of the ITS-ECO revealed CuO ENMs to elicit cytotoxic effects on lysosomes of hemocytes of mussels, with the hazard potential CuO PEG > CuO COOH > CuO NH3 > CuO core, whereas TiO2 ENMs were not cytotoxic. Genotoxicity in hemocytes as well as gill cells of mussels following in vivo exposure (48 h) to CuO ENMs was also seen. Longer in vivo exposures in Tier 2 (48 h-21 days) revealed subacute and chronic oxidative effects for both CuO and TiO2 ENMs, in some cases leading to lipid peroxidation (core TiO2 ENMs). In Tier 3 bioaccumulation studies, distinct patterns of uptake for Cu (predominantly in gills) and Ti (predominantly in digestive glands) and between the different core and coated ENMs were found. Clear NM-specific and coating-dependent effects on hazard and fate were seen. Overall, using a tiered testing approach, the ITS-ECO was able to differentiate the hazard (acute, subacute, and chronic effects) posed by ENMs of different compositions and coatings and to provide information on fate for environmental risk assessment of these ENMs. Environ Toxicol Chem 2022;41:1390-1406. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

An integrated approach to testing and assessment of high aspect ratio nanomaterials and its application for grouping based on a common mesothelioma hazard.

Here we describe the development of an Integrated Approach to Testing and Assessment (IATA) to support the grouping of different types (nanoforms; NFs) of High Aspect Ratio Nanomaterials (HARNs), based on their potential to cause mesothelioma. Hazards posed by the inhalation of HARNs are of particular concern as they exhibit physical characteristics similar to pathogenic asbestos fibres. The approach for grouping HARNs presented here is part of a framework to provide guidance and tools to group similar NFs and aims to reduce the need to assess toxicity on a case-by-case basis. The approach to grouping is hypothesis-driven, in which the hypothesis is based on scientific evidence linking critical physicochemical descriptors for NFs to defined fate/toxicokinetic and hazard outcomes. The HARN IATA prompts users to address relevant questions (at decision nodes; DNs) regarding the morphology, biopersistence and inflammatory potential of the HARNs under investigation to provide the necessary evidence to accept or reject the grouping hypothesis. Each DN in the IATA is addressed in a tiered manner, using data from simple in vitro or in silico methods in the lowest tier or from in vivo approaches in the highest tier. For these proposed methods we provide justification for the critical descriptors and thresholds that allow grouping decisions to be made. Application of the IATA allows the user to selectively identify HARNs which may pose a mesothelioma hazard, as demonstrated through a literature-based case study. By promoting the use of alternative, non-rodent approaches such as in silico modelling, in vitro and cell-free tests in the initial tiers, the IATA testing strategy streamlines information gathering at all stages of innovation through to regulatory risk assessment while reducing the ethical, time and economic burden of testing.

Toward sustainable environmental quality: Priority research questions for Europe.

The United Nations' Sustainable Development Goals have been established to end poverty, protect the planet, and ensure prosperity for all. Delivery of the Sustainable Development Goals will require a healthy and productive environment. An understanding of the impacts of chemicals which can negatively impact environmental health is therefore essential to the delivery of the Sustainable Development Goals. However, current research on and regulation of chemicals in the environment tend to take a simplistic view and do not account for the complexity of the real world, which inhibits the way we manage chemicals. There is therefore an urgent need for a step change in the way we study and communicate the impacts and control of chemicals in the natural environment. To do this requires the major research questions to be identified so that resources are focused on questions that really matter. We present the findings of a horizon-scanning exercise to identify research priorities of the European environmental science community around chemicals in the environment. Using the key questions approach, we identified 22 questions of priority. These questions covered overarching questions about which chemicals we should be most concerned about and where, impacts of global megatrends, protection goals, and sustainability of chemicals; the development and parameterization of assessment and management frameworks; and mechanisms to maximize the impact of the research. The research questions identified provide a first-step in the path forward for the research, regulatory, and business communities to better assess and manage chemicals in the natural environment. Environ Toxicol Chem 2018;37:2281-2295. © 2018 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals, Inc. on behalf of SETAC.

Regulatory ecotoxicity testing of nanomaterials - proposed modifications of OECD test guidelines based on laboratory experience with silver and titanium dioxide nanoparticles.

Regulatory ecotoxicity testing of chemicals is of societal importance and a large effort is undertaken at the OECD to ensure that OECD test guidelines (TGs) for nanomaterials (NMs) are available. Significant progress to support the adaptation of selected TGs to NMs was achieved in the context of the project MARINA ( http://www.marina-fp7.eu/ ) funded within the 7th European Framework Program. Eight OECD TGs were adapted based on the testing of at least one ion-releasing NM (Ag) and two inert NMs (TiO2). With the materials applied, two main variants of NMs (ion releasing vs. inert NMs) were addressed. As the modifications of the test guidelines refer to general test topics (e.g. test duration or measuring principle), we assume that the described approaches and modifications will be suitable for the testing of further NMs with other chemical compositions. Firm proposals for modification of protocols with scientific justification(s) are presented for the following tests: growth inhibition using the green algae Raphidocelis subcapitata (formerly: Pseudokirchneriella subcapitata; TG 201), acute toxicity with the crustacean Daphnia magna (TG 202), development toxicity with the fish Danio rerio (TG 210), reproduction of the sediment-living worm Lumbriculus variegatus (TG 225), activity of soil microflora (TGs 216, 217), and reproduction of the invertebrates (Enchytraeus crypticus, Eisenia fetida, TGs 220, 222). Additionally, test descriptions for two further test systems (root elongation of plants in hydroponic culture; test on fish cells) are presented. Ecotoxicological data obtained with the modified test guidelines for TiO2 NMs and Ag NM and detailed method descriptions are available.

A Multilaboratory Toxicological Assessment of a Panel of 10 Engineered Nanomaterials to Human Health--ENPRA Project--The Highlights, Limitations, and Current and Future Challenges.

ENPRA was one of the earlier multidisciplinary European Commission FP7-funded projects aiming to evaluate the risks associated with nanomaterial (NM) exposure on human health across pulmonary, cardiovascular, hepatic, renal, and developmental systems. The outputs from this project have formed the basis of this review. A retrospective interpretation of the findings across a wide range of in vitro and in vivo studies was performed to identify the main highlights from the project. In particular, focus was placed on informing what advances were made in the hazard assessment of NM, as well as offering some suggestions on the future of "nanotoxicology research" based on these observations, shortcomings, and lessons learned from the project. A number of issues related to the hazard assessment of NM are discussed in detail and include use of appropriate NM for nanotoxicology investigations; characterization and dispersion of NM; use of appropriate doses for all related investigations; need for the correct choice of experimental models for risk assessment purposes; and full understanding of the test systems and correct interpretation of data generated from in vitro and in vivo systems. It is hoped that this review may assist in providing information in the implementation of guidelines, model systems, validation of assessment methodology, and integrated testing approaches for risk assessment of NM. It is vital to learn from ongoing and/or completed studies to avoid unnecessary duplication and offer suggestions that might improve different aspects of experimental design.

The MARINA Risk Assessment Strategy: A Flexible Strategy for Efficient Information Collection and Risk Assessment of Nanomaterials.

An engineered nanomaterial (ENM) may actually consist of a population of primary particles, aggregates and agglomerates of various sizes. Furthermore, their physico-chemical characteristics may change during the various life-cycle stages. It will probably not be feasible to test all varieties of all ENMs for possible health and environmental risks. There is therefore a need to further develop the approaches for risk assessment of ENMs. Within the EU FP7 project Managing Risks of Nanoparticles (MARINA) a two-phase risk assessment strategy has been developed. In Phase 1 (Problem framing) a base set of information is considered, relevant exposure scenarios (RESs) are identified and the scope for Phase 2 (Risk assessment) is established. The relevance of an RES is indicated by information on exposure, fate/kinetics and/or hazard; these three domains are included as separate pillars that contain specific tools. Phase 2 consists of an iterative process of risk characterization, identification of data needs and integrated collection and evaluation of data on the three domains, until sufficient information is obtained to conclude on possible risks in a RES. Only data are generated that are considered to be needed for the purpose of risk assessment. A fourth pillar, risk characterization, is defined and it contains risk assessment tools. This strategy describes a flexible and efficient approach for data collection and risk assessment which is essential to ensure safety of ENMs. Further developments are needed to provide guidance and make the MARINA Risk Assessment Strategy operational. Case studies will be needed to refine the strategy.

Nanopesticides: guiding principles for regulatory evaluation of environmental risks.

Nanopesticides or nano plant protection products represent an emerging technological development that, in relation to pesticide use, could offer a range of benefits including increased efficacy, durability, and a reduction in the amounts of active ingredients that need to be used. A number of formulation types have been suggested including emulsions (e.g., nanoemulsions), nanocapsules (e.g., with polymers), and products containing pristine engineered nanoparticles, such as metals, metal oxides, and nanoclays. The increasing interest in the use of nanopesticides raises questions as to how to assess the environmental risk of these materials for regulatory purposes. Here, the current approaches for environmental risk assessment of pesticides are reviewed and the question of whether these approaches are fit for purpose for use on nanopesticides is addressed. Potential adaptations to existing environmental risk assessment tests and procedures for use with nanopesticides are discussed, addressing aspects such as analysis and characterization, environmental fate and exposure assessment, uptake by biota, ecotoxicity, and risk assessment of nanopesticides in aquatic and terrestrial ecosystems. Throughout, the main focus is on assessing whether the presence of the nanoformulation introduces potential differences relative to the conventional active ingredients. The proposed changes in the test methodology, research priorities, and recommendations would facilitate the development of regulatory approaches and a regulatory framework for nanopesticides.

Concern-driven integrated approaches to nanomaterial testing and assessment--report of the NanoSafety Cluster Working Group 10.

Bringing together topic-related European Union (EU)-funded projects, the so-called "NanoSafety Cluster" aims at identifying key areas for further research on risk assessment procedures for nanomaterials (NM). The outcome of NanoSafety Cluster Working Group 10, this commentary presents a vision for concern-driven integrated approaches for the (eco-)toxicological testing and assessment (IATA) of NM. Such approaches should start out by determining concerns, i.e., specific information needs for a given NM based on realistic exposure scenarios. Recognised concerns can be addressed in a set of tiers using standardised protocols for NM preparation and testing. Tier 1 includes determining physico-chemical properties, non-testing (e.g., structure-activity relationships) and evaluating existing data. In tier 2, a limited set of in vitro and in vivo tests are performed that can either indicate that the risk of the specific concern is sufficiently known or indicate the need for further testing, including details for such testing. Ecotoxicological testing begins with representative test organisms followed by complex test systems. After each tier, it is evaluated whether the information gained permits assessing the safety of the NM so that further testing can be waived. By effectively exploiting all available information, IATA allow accelerating the risk assessment process and reducing testing costs and animal use (in line with the 3Rs principle implemented in EU Directive 2010/63/EU). Combining material properties, exposure, biokinetics and hazard data, information gained with IATA can be used to recognise groups of NM based upon similar modes of action. Grouping of substances in return should form integral part of the IATA themselves.

Minimal analytical characterization of engineered nanomaterials needed for hazard assessment in biological matrices.

This paper presents the outcomes from a workshop of the European Network on the Health and Environmental Impact of Nanomaterials (NanoImpactNet). During the workshop, 45 experts in the field of safety assessment of engineered nanomaterials addressed the need to systematically study sets of engineered nanomaterials with specific metrics to generate a data set which would allow the establishment of dose-response relations. The group concluded that international cooperation and worldwide standardization of terminology, reference materials and protocols are needed to make progress in establishing lists of essential metrics. High quality data necessitates the development of harmonized study approaches and adequate reporting of data. Priority metrics can only be based on well-characterized dose-response relations derived from the systematic study of the bio-kinetics and bio-interactions of nanomaterials at both organism and (sub)-cellular levels. In addition, increased effort is needed to develop and validate analytical methods to determine these metrics in a complex matrix.

The importance of life cycle concepts for the development of safe nanoproducts.

Whilst the global players in industry are rapidly moving forward to take advantage of the new opportunities and prospects offered by nanotechnologies, it is imperative that such developments take place in a safe and sustainable manner. The increasing use of engineered nanomaterials (ENMs) in consumer products has raised certain concerns over their safety to human health and the environment. There are currently a number of major uncertainties and knowledge gaps in regard to behavior, chemical and biological interactions and toxicological properties of ENMs. As dealing with these uncertainties will require the generation of new basic knowledge, it is unlikely that they will be resolved in the immediate future. One has to consider the whole life cycle of nanoproducts to ensure that possible impacts can be systematically discovered. For example, life cycle assessment (LCA) - a formalized life cycle concept - may be used to assess the relative environmental sustainability performance of nanoproducts in comparison with their conventional equivalents. Other less formalized life cycle concepts in the framework of prospective technology assessment may uncover further detailed and prospective knowledge for human and environmental exposure to ENMs during the life cycle of nanoproducts. They systematically reveal impacts such as cross product contamination or dissipation of scarce materials among others. The combination of different life cycle concepts with the evolving knowledge from toxicology and risk assessment can mitigate uncertainties and can provide an early basis for informed decision making by the industry and regulators.