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Mechanistic modelling of Critical Target Tissue Residues based on substance properties, species characteristics and external effect concentrations

Final Report Summary - MEMOCTR (Mechanistic modelling of Critical Target Tissue Residues based on substance properties, species characteristics and external effect concentrations)

A key challenge facing ecotoxicology is the need to assess the risk of thousands of substances, for a wide range of species, with high accuracy and ecological relevance. Toxicokinetic/toxicodynamic (TKTD) models are advocated as essential tools to address this challenge, but are currently underutilized. This partly results from model-parameterization difficulties: internal target site concentrations are difficult to establish experimentally and model parameters are presently inferred for each toxicant-species combination specifically by measuring both the time-course of internal toxicant concentration (toxicokinetics) and the time-course of toxic response (toxicodynamics). TKTD modelling can significantly advance if both TK and TD parameters are quantitatively related to key chemical properties and key species characteristics. At present, such relationships are relatively well-established for toxicokinetics in the form of mechanistic bioaccumulation models, but analogous models are not yet available for toxicodynamics. We want to further advance TKTD modelling by quantitatively linking relevant TD parameters to chemical properties and species characteristics. We focus on acute toxicity prediction of metal exposure in freshwater aquatic organisms. The overall aim is to develop mechanistic TKTD models for acute toxicity prediction of metals in various freshwater organisms using metal-properties, species-characteristics and external effect concentrations. Sub-objectives are to: i) to develop species-specific quantitative structure activity relationships (QSARs) that relate external toxicity metrics (LC50) of various metals to a metal-specific property, ii) To quantitatively relate external toxicity metrics of selected metals to a species characteristic, iii) to characterize and quantify key biochemical and physiological processes that determine the biological response on a molecular and individual level and integrate those in a toxicodynamic model, iv) to develop a new mechanistic TKTD model by integrating the relevant biochemical and physiological processes into a TKTD model framework for survival of organisms, and v) to test the developed TKTD model with empirical data of sub-lethal effects (e.g. target enzyme inhibition, sub-lethal physiological responses) and external acute toxicity data for various aquatic species. All required data are collected from peer-reviewed literature and relevant databases such as US EPA’s Aquire.
Our results show that acute toxicity is significantly related to the metal-specific covalent index for various species, incl. rainbow trout, Daphnia magna and the pond snail. Regressions were developed for 12 different species in total. All regressions were significant (p < 0.05) and r2 ranged between 0.53 and 0.90. Interestingly, the slopes of the LC50-Χ2 mr regression lines are remarkably similar across species, with slopes ranging from -1.08 to -1.59. This suggests that the mode-of-action is similar for a range of freshwater aquatic organisms. The implication for TKTD modelling is that the covalent index is an important metal-specific property that should be considered. Additionally, the results suggest thata common TKTD model framework may be applied to a range of freshwater organisms. Our results furthermore show that acute toxicity of both copper (Cu) and silver (Ag) is significantly related to species weight. It is found that LC50s relate to species weight to the power 0.21 (for Ag) and 0.23 (for Cu) (Fig. 1), which is remarkably close to the well-established allometric scaling exponent of ¼ for metabolic rates. Size is thus an important factor to consider when developing a TKTD model for acute toxicity of Ag and Cu. In addition, this suggest that an important rate-limiting step in the adverse outcome pathway is related to species size.

a. Silver (Ag)
b. Copper (Cu)

Figure 1. Acute silver and copper toxicity (48h/96h LC50) of various freshwater aquatic organisms plotted against species weight (W). The dotted line represents the regression analysis of LogLC50 vs. LogW. The error bars represent 95% confidence intervals.
We proceeded to develop a TKTD model for Ag and Cu. Both Cu and Ag elicit their acute toxic action by inhibiting Na+/K+-ATPase enzymes which are located at the basolateral membrane of the gill.
Inhibition of this enzyme results in a reduced sodium influx and a fall in whole-body sodium concentrations. Rainbow trout were observed to die when they had lossed approximately 30% of their original exchangeable sodium concentration and it has been suggested that this ‘critical Na loss’ threshold is applicable to other freshwater aquatic organisms as well (e.g. Paquin et al. 2002).
Additionally, it has been suggested that the observed scaling of LC50s to species size is a result of the scaling of sodium efflux rates to species size (e.g. Grosell et al. 2000). We developed a toxicodynamic model that quantifies the reduction in whole-body sodium concentration over time as a function of the metal-specific inhibition of the target site over time. A sodium mass-balance model was developed that quantifies sodium kinetics in various freshwater organisms as a function of species size. Furthermore, we collected data on %survival of aquatic organisms and %sodium loss, to determine whether the ‘critical sodium threshold’ is applicable to other organisms than the rainbow trout. First results show that survival of aquatic organisms is significantly related to sodium loss (r2 = 0.86). However, sodium loss kinetics are in itself too fast to explain the observed scaling of external acute toxicity data with species weight. A comparison of the predicted fraction of non-functional target enzymes with empirical data, suggests that the model overestimates target enzyme inhibition. Further research thus focusses on improving the toxicodynamic model and extending the developed TKTD framework to predict acute toxicity of other metals (e.g. Zn, Cd, Hg, Pb, Ni) to freshwater aquatic organisms. Development of such mechanistic TKTD models is useful for environmental risk assessment as quantitatively linking both TK and TD model parameters to metal properties and species characteristics allows for model application to untested metals and to untested species.

References
Paquin PR, Zoltay V, Winfield RP, Wu KB, Mathew R, Santore RC, DiToro DM. Extension of the Biotic Ligand Model for acute toxicity to a physiologically-based model of the survival time of rainbow trout (Oncorhynchus mykiss) exposed to silver. Comp. Biochem. Physiol. Part C. 2002, 133, 305-343
Grosell M, Nielsen C, Bianchini A. Sodium turnover rate determines sensitivity to acute copper and silver exposure in freshwater animals. Comp. Biochem. Physiol. Part C: Toxicol. Pharmacol. 2002, 133 (1-2), 287-303