Kristian Syberg

On the Use of Mixture Toxicity Assessment in REACH and the Water Framework Directive: A Review

ABSTRACT This review seeks to connect the scientific theory of mixture toxicity to its implementation within different regulatory frameworks. The aim is to demonstrate how mixture toxicity assessment can be more thoroughly integrated into the European chemical regulations, REACH, and the Water Framework Directive (WFD), using the experiences gained through other regulatory frameworks. The article consists of (1) an examination of the scientific underpinnings of the common mixture toxicity assessment methods; (2) a discussion of how these methods have been used in regulatory frameworks; and (3) a discussion of how the methods could be applied within REACH and the WFD. It is concluded that concentration addition should be applied as a default model for mixture toxicity assessment. Furthermore, it is concluded that REACH and the WFD only include mixture toxicity assessments in specific situations. However, it is shown that it is scientifically feasible and regulatorally practicable to integrate a more holistic mixture toxicity approach into both legislations. In this connection, the experience gained from the U.S. frameworks on mixture toxicity assessment could be useful. The construction of a database that includes data on chemicals in the European environment could be used for mixture toxicity assessment of the chemicals with individual PEC/PNECs > 0.1.

Assessing and managing multiple risks in a changing world – The Roskilde recommendations

Roskilde University (Denmark) hosted a November 2015 workshop, Environmental Risk-Assessing and Managing Multiple Risks in a Changing World. This Focus article presents the consensus recommendations of 30 attendees from 9 countries regarding implementation of a common currency (ecosystem services) for holistic environmental risk assessment and management; improvements to risk assessment and management in a complex, human-modified, and changing world; appropriate development of protection goals in a 2-stage process; dealing with societal issues; risk-management information needs; conducting risk assessment of risk management; and development of adaptive and flexible regulatory systems. The authors encourage both cross-disciplinary and interdisciplinary approaches to address their 10 recommendations: 1) adopt ecosystem services as a common currency for risk assessment and management; 2) consider cumulative stressors (chemical and nonchemical) and determine which dominate to best manage and restore ecosystem services; 3) fully integrate risk managers and communities of interest into the risk-assessment process; 4) fully integrate risk assessors and communities of interest into the risk-management process; 5) consider socioeconomics and increased transparency in both risk assessment and risk management; 6) recognize the ethical rights of humans and ecosystems to an adequate level of protection; 7) determine relevant reference conditions and the proper ecological context for assessments in human-modified systems; 8) assess risks and benefits to humans and the ecosystem and consider unintended consequences of management actions; 9) avoid excessive conservatism or possible underprotection resulting from sole reliance on binary, numerical benchmarks; and 10) develop adaptive risk-management and regulatory goals based on ranges of uncertainty. Environ Toxicol Chem 2017;36:7-16.

Comet assay on tetraploid yeast cells.

Tetraploid yeast cells (Saccharomyces cerevisiae) were used in the comet assay with the intention of developing a new, fast and easy assay for detecting environmental genotoxic agents without using higher organisms. Two DNA-damaging chemicals, H(2)O(2) and acrylamide, together with wastewater from three municipal treatment plants were tested for their effect on the yeast-cell DNA. The main problem with using yeast in the comet assay is the necessity to degrade the cell wall. This was achieved by using Zymolase 100 T twice during the procedure, since Zymolase 20 T did not open the cell wall. Analytical problems that arose due to the small amount of DNA in the yeast nuclei in haploid and diploid cells, which contain 13 Mbp and 26 Mbp DNA per cell, respectively, were solved by using tetraploid yeast cells (52 Mbp) instead. DNA damage was shown after exposure to H(2)O(2) and acrylamide. The lowest dose causing significant DNA damage was 20 microM for H(2)O(2) and 200mg/l for acrylamide. Tertiary-treated wastewater from the outlets of three municipal wastewater-treatment plants was tested, but did not cause DNA damage. Even though it is possible to produce comets with tetraploid yeast cells, the amount of DNA is likely too small for a proper comet assay.

From macro- to microplastics - Analysis of EU regulation along the life cycle of plastic bags ☆

Plastic pollution and its environmental effects has received global attention the recent years. However, limited attention has so far been directed towards how plastics are regulated in a life cycle perspective and how regulatory gaps can be addressed in order to limit and prevent environmental exposure and hazards of macro- and microplastics. In this paper, we map European regulation taking outset in the life cycle perspective of plastic carrier bags: from plastic bag production to when it enters the environment. Relevant regulatory frameworks, directives and authorities along the life cycle are identified and their role in regulation of plastics is discussed. Most important regulations were identified as: the EU chemical Regulation, the Packaging and Packaging Waste Directive including the amending Directive regarding regulation of the consumption of lightweight plastic carrier bags, the Waste Framework Directive and the Directive on the Landfill of Waste. The main gaps identified relate to lack of clear definitions of categories of polymers, unambitious recycling rates and lack of consideration of macro- and microplastics in key pieces of legislation. We recommend that polymers are categorized according to whether they are polymers with the same monomer constituents (homopolymers) or with different monomer constituents (copolymers) and that polymers are no longer exempt from registration and evaluation under REACH. Plastics should furthermore have the same high level of monitoring and reporting requirements as hazardous waste involving stricter requirements to labelling, recordkeeping, monitoring and control over the whole lifecycle. Finally, we recommend that more ambitious recycle and recovery targets are set across the EU. Regulation of the consumption of lightweight plastic carrier bags should also apply to heavyweight plastic carrier bags. Last, the Marine and Water Framework Directives should specifically address plastic waste affecting water quality.

Genotoxic potential of two herbicides and their active ingredients assessed with comet assay on a fish cell line, epithelioma papillosum cyprini (EPC).

The aim of this study was to optimize the epithelioma papillosum cyprini (EPC) cell line handling procedure for the comet assay to investigate the genotoxic potential of widely used pesticides. The effects of various media and handling of the EPC cell line were examined. Results indicated that avoiding trypsin to detach cells led to lower level of DNA damage in the negative control. Further, two commonly used herbicides (Dezormon and Optica trio) and their four active ingredients (4-chloro-o-tolyloxyacetic acid, 2,4-dichlorophenoxyacetic acid, 2-(4-chloro-2-methylphenoxy)propionic acid, 2-(2,4-dichlorophenoxy)propionic acid) individually and in a ternary mixture were examined with the comet assay. Data showed that among the active ingredients only 2,4-D and MCPA induced DNA damage, while both herbicides were genotoxic at high concentrations.

Microplastic potentiates triclosan toxicity to the marine copepod Acartia tonsa (Dana)

ABSTRACT Microplastics (MP) are contaminants of environmental concern partly due to plastics ability to sorb and transport hydrophobic organic contaminants (HOC). The importance of this “vector effect” is currently being debated in the scientific community. This debate largely ignores that the co-exposures of MP and HOC are mixtures of hazardous agents, which can be addressed from a mixture toxicity perspective. In this study, mixture effects of polyethylene microbeads (MP) and triclosan (TCS) (a commonly used antibacterial agent in cosmetics) were assessed on the marine copepod Acartia tonsa. Data indicated that MP potentiate the toxicity of TCS, illustrating the importance of understanding the mixture interaction between plastics and HOC when addressing the environmental importance of the vector effect.

Microplastics in Inland African Waters: Presence, Sources, and Fate

As the birthplace of our species, the African continent holds a unique place in human history. Upon entering a new epoch, the Anthropocene defined by human-driven influences on earth systems, and with the recognition that plastic pollution is one of the hallmarks of this new age, remarkably little is known about the presence, sources, and fate of plastics (and microplastics (MPs)) within African waters. Research in marine regions, most notably around the coast of South Africa, describes the occurrence of MPs in seabirds and fish species. More recently environmental sampling studies in the same area have quantified plastics in both the water column and sediments. However, despite Africa containing some of the largest and deepest of the world’s freshwater lakes, including Lakes Victoria and Tanganyika as part of the African Great Lakes system, and notable freshwater rivers, such as the River Congo and the Nile, the extent of MPs within the inland waters remains largely unreported. In the only study to date to describe MP pollution in the African Great Lakes, a variety of polymers, including polyethylene, polypropylene, and silicone rubber, were recovered from the gastrointestinal tracts of Nile perch (Lates niloticus) and Nile tilapia (Oreochromis niloticus) fished from Lake Victoria. The likely sources of these plastics were considered to be human activities linked to fishing and tourism, and urban waste. In this chapter we discuss the need for research focus on MPs in Africa and how what has been described in the coastal regions and other freshwater environments can be applied to inland African waters. The aforementioned study in Lake Victoria is used to exemplify how small-scale investigations can provide early indications of MP pollution. Lastly we discuss the current challenges and future needs of MP research in African freshwaters.

Toward a conceptual approach for assessing risks from chemical mixtures and other stressors to coastal ecosystem services.

Growth of human populations and increased human activity, particularly in coastal areas, increase pressure on coastal ecosystems and the ecosystem services (ES) they provide. As a means toward being able to assess the impact of multiple stressors on ES, in the present study we propose an 8-step conceptual approach for assessing effects of chemical mixtures and other stressors on ES in coastal areas: step A, identify the relevant problems and policy aims; step B, identify temporal and spatial boundaries; step C, identify relevant ES; step D, identify relevant stressors (e.g., chemicals); step E, translate impacts into ES units; step F, assess cumulative risk in ES units; step G, rank stressors based on their contribution to adverse effects on ES; and step H, implement regulation and management as appropriate and necessary. Two illustrative case studies (Swedish coastal waters and a coastal lagoon in Costa Rica) are provided; one focuses on chemicals that affect human food supply and the other addresses pesticide runoff and trade-offs among ES. The 2 cases are used to highlight challenges of such risk assessments, including use of standardized versus ES-relevant test species, data completeness, and trade-offs among ES. Lessons learned from the 2 case studies are discussed in relation to environmental risk assessment and management of chemical mixtures. Integr Environ Assess Manag 2017;13:376-386.

Mixture Genotoxicity of 2,4-Dichlorophenoxyacetic Acid, Acrylamide, and Maleic Hydrazide on Human Caco-2 Cells Assessed with Comet Assay

Assessment of genotoxic properties of chemicals is mainly conducted only for single chemicals, without taking mixture genotoxic effects into consideration. The current study assessed mixture effects of the three known genotoxic chemicals, 2,4-dichlorophenoxyacetic acid (2,4-D), acrylamide (AA), and maleic hydrazide (MH), in an experiment with a fixed ratio design setup. The genotoxic effects were assessed with the single-cell gel electrophoresis assay (comet assay) for both single chemicals and the ternary mixture. The concentration ranges used were 0–1.4, 0–20, and 0–37.7 mM for 2,4-D, AA, and MH, respectively. Mixture toxicity was tested with a fixed ratio design at a 10:23:77% ratio for 2.4-D:AA:MH. Results indicated that the three chemicals yielded a synergistic mixture effect. It is not clear which mechanisms are responsible for this interaction. A few possible interactions are discussed, but further investigations including in vivo studies are needed to clarify how important these more-than-additive effects are for risk assessment.

Are Standardized Test Guidelines Adequate for Assessing Waterborne Particulate Contaminants

O the first step in identifying the ecotoxicological risk and producing regulatory guidance for new chemical substances is the performance of standardized toxicity tests. Protocols described within the test guidelines of the Organisation for Economic Cooperation and Development’s “Effects on Biotic Systems” outline methodologies to determine toxicities, defined by specific end points and model organisms within the aquatic environment. These procedures are equipped for the assessement of chemical solutes, but critical deficiencies in standardized test guidelines are increasingly apparent in the determination of toxicities resulting from the dispersion of particulates in water. Our aim is to highlight these shortcomings and promote discussion of how testing methods could be adapted to an increasingly common form of contaminant. In the past decade two of the most prominent research areas in ecotoxicology have been engineered nanomaterials (ENMs, one dimension <100 nm in size (see Virtual Issue on Environmental Nanotoxicology (http://pubs.acs.org/page/vi/ enviro_nanotoxicology.html)) and microplastics (MPs; synthetic organic polymers produced (primary MPs) or degraded (secondary MPs) to <5 mm in size). The emerging issue of nanoplastics (i.e., plastics in the nanosize range) continues this focus on particulates. The inadequacies of standardized testing have been previously highlighted with regard to specific particulates (ENMs). Although some of these considerations could be generalized to many anthropogenic waterborne particulates, a reassessment of standardized approaches and their application to particulates is now timely. We describe five specific limitations of standardized test methods that, while designed for chemical solutes, fail to capture the intricies of particulate exposures: