Three-tiered approach for standard information requirements for polymers requiring registration under REACH.

Polymers are a very large class of chemicals comprising often complex molecules with multiple functions used in everyday products. The EU Commission is seeking to develop environmental and human health standard information requirements (SIRs) for man-made polymers requiring registration (PRR) under a revised Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH) Regulation. Conventional risk assessment approaches currently used for small molecules may not apply to most polymers. Therefore, we propose a conceptual three-tiered regulatory approach for data generation to assess individual and groups of polymers requiring registration (PRR). A key element is the grouping of polymers according to chemistry, physico-chemical properties and hazard similarity. The limited bioavailability of many polymers is a prominent difference to many small molecules and is a key consideration of the proposed approach. Methods assessing potential for systemic bioavailability are integral to Tier 1. Decisions for further studies are based on considerations of properties and effects, combined with systemic bioavailability and use and exposure considerations. For many PRRs, Tier 1 data on hazard, use and exposure will likely be sufficient for achieving the protection goals of REACH. Vertebrate animal studies in Tiers 2 and 3 can be limited to targeted testing. The outlined approach aims to make use of current best scientific evidence and to reduce animal testing whilst providing data for an adequate level of protection.

Bioconcentration Assessment in Fish Based on In Vitro Intrinsic Clearance: Predictivity of an Empirical Model Compared to In Vitro-In Vivo Extrapolation Models.

To estimate the bioconcentration factor (BCF), the in vitro intrinsic clearance (CLIN VITRO,INT) from rainbow trout liver S9 fractions (RT-S9) can be applied to in vitro-in vivo extrapolation (IVIVE) models, yet uncertainties remain in model parameterization. An alternative model approach is evaluated: a regression model was built in the form log BCF = a × log Kow + b × log CLIN VITRO,INT. The coefficients a and b were fitted based on a training set of 40 chemicals. A high robustness of the coefficients and good accuracy of BCF prediction were found on independent datasets of neutral organic chemicals (measured log Kow 3.3-6.2). BCF predictions were similar to or in better agreement with in vivo BCFs compared to IVIVE models (2.4- to 2.9- vs 2.8- to 3.6-fold misprediction) for training and test sets. Species-matched models (trout, carp) did not result in improvements. This study presents the largest dataset on CLIN VITRO,INT and BCFs to assess predictivity of the RT-S9 assay. The robustness of the regression statistics on different datasets and the high statistical weight of the CLIN VITRO,INT term illustrate the predictive power of the RT-S9 assay as an important step toward regulatory acceptance to replace animal experiments.

Mobility in the context of exposure-based assessment of chemicals for drinking water resource protection.

In order to protect European Union (EU) drinking water resources from chemical contamination, criteria for identifying persistent, mobile, and toxic (PMT) chemicals and very persistent and very mobile (vPvM) chemicals under the EU REACH Regulation were proposed by the German Environment Agency (Umweltbundesamt-UBA). Additionally, new hazard classes for PMT and vPvM substances in the revised EU classification, labeling, and packaging (CLP Regulation) are intended. Therefore, a reliable approach in the identification of potential drinking water resource contaminants is needed. The scientific basis of the property-based PMT/vPvM criteria, focusing on mobility, which dictates the migration of chemical drinking water sources, was evaluated, and a critical analysis of the deviation of sorption metrics from simple behavior was carried out. Based on our evaluation, a Koc may be used for nonionic substances on a screening level only, requiring a higher tier assessment. It is considered inappropriate for hydrophilic and ionizable chemicals, particularly for soils with low organic carbon contents. The nonextractable residue formation is complex and not well understood but remains significant in limiting the mobility of chemicals through soils and sediments. In order to inform the EU commission's work on the introduction of new hazard classes for PMT and vPvM substances into the European legislation, the derivation of a tiered approach is proposed, which utilizes the weight of evidence available, with adoption of appropriate higher tier models commensurate with the nature of the substance and the data available. Integr Environ Assess Manag 2023;19:775-791. © 2022 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC).

Examining Uncertainty in In Vitro-In Vivo Extrapolation Applied in Fish Bioconcentration Models.

In vitro biotransformation rates were determined for 30 chemicals, mostly fragrance ingredients, using trout liver S9 fractions (RT-S9) and incorporated into in vitro-in vivo extrapolation (IVIVE) models to predict bioconcentration factors (BCFs). Predicted BCFs were compared against empirical BCFs to explore potential major uncertainties involved in the in vitro methods and IVIVE models: (i) in vitro chemical test concentrations; (ii) different gill uptake rate constant calculations (k1); (iii) protein binding (different calculations and measurement of the fraction of unbound chemical, fU); (iv) species differences; and (v) extrahepatic biotransformation. Predicted BCFs were within 0.5 log units for 44% of the chemicals compared to empirical BCFs, whereas 56% were overpredicted by >0.5 log units. This trend of overprediction was reduced by alternative k1 calculations to 32% of chemicals being overpredicted. Moreover, hepatic in vitro rates scaled to whole body biotransformation rates (kB) were compared against in vivo kB estimates. In vivo kB was underestimated for 79% of the chemicals. Neither lowering the test concentration, nor incorporation of new measured fU values, nor species matching avoided the tendency to overpredict BCFs indicating that further improvements to the IVIVE models are needed or extrahepatic biotransformation plays an underestimated role.

PeBiToSens™: A Platform for PBT Screening of Fragrance Ingredients Without Animal Testing.

The determination of persistence (P), bioaccumulation (B) and toxicity (T) plays a central role in the environmental assessment of chemicals. Persistence is typically evaluated via standard microbial biodegradation tests. Bioaccumulation refers to the accumulation of chemicals in organisms and is usually assessed in fish exposed to the test chemical. Toxicity is determined at three trophic levels, with fish toxicity as the highest trophic level assessed. Thus, animal tests are classically needed for both B and T assessment. In vitro systems based on fish liver cells or liver S9 fractions ('RT-S9 assay') have been recently adopted by OECD to measure the biotransformation rates for the chemicals for B assessment. Biotransformation drives clearance from the body and reduces bioaccumulation. For T assessment, an assay based on in vitro toxicity on fish gill cells has been established ('RTgill-W1 assay'). Here we summarize our findings indicating that these tests are highly predictive for fragrance ingredients, and show with two case studies of our latest new registered substances how we apply these tests in particular during development and also for chemical registration. This platform of tests (PeBiToSens™) could fully replace animal tests in ecotoxicological assessment and is key in the Givaudan Safe by Design™ approach to develop safer and environmentally compatible novel fragrance ingredients.

Accurate prediction of acute fish toxicity of fragrance chemicals with the RTgill-W1 cell assay.

Testing for acute fish toxicity is an integral part of the environmental safety assessment of chemicals. A true replacement of primary fish tissue was recently proposed using cell viability in a fish gill cell line (RTgill-W1) as a means of predicting acute toxicity, showing good predictivity on 35 chemicals. To promote regulatory acceptance, the predictivity and applicability domain of novel tests need to be carefully evaluated on chemicals with existing high-quality in vivo data. We applied the RTgill-W1 cell assay to 38 fragrance chemicals with a wide range of both physicochemical properties and median lethal concentration (LC50) values and representing a diverse range of chemistries. A strong correlation (R2 = 0.90-0.94) between the logarithmic in vivo LC50 values, based on fish mortality, and the logarithmic in vitro median effect concentration (EC50) values based on cell viability was observed. A leave-one-out analysis illustrates a median under-/overprediction from in vitro EC50 values to in vivo LC50 values by a factor of 1.5. This assay offers a simple, accurate, and reliable alternative to in vivo acute fish toxicity testing for chemicals, presumably acting mainly by a narcotic mode of action. Furthermore, the present study provides validation of the predictivity of the RTgill-W1 assay on a completely independent set of chemicals that had not been previously tested and indicates that fragrance chemicals are clearly within the applicability domain. Environ Toxicol Chem 2018;37:931-941. © 2017 SETAC.