In Vitro Assessment of Modes of Toxic Action of Pharmaceuticals in Aquatic Life

Abstract

An ecotoxicological test battery based on a mode-of-action approach was designed and applied to the hazard identification and classification of modes of action of six pharmaceuticals (carbamazepine, diclofenac, ethinyl estradiol, ibuprofen, propranolol, and sulfamethoxazole). The rationale behind the design of the battery was to cover the relevant interactions that a compound may have with biological targets. It is thus not comprehensive but contains representative examples of each category of mode of toxic action including nonspecific, specific, and reactive toxicity. The test battery consists of one test system for nonspecific toxicity (baseline toxicity or narcosis), two test systems for specific effects, and two test systems for reactive toxicity. The baseline toxicity was quantified with the Kinspec test, which detects membrane leakage via measurements of membrane potential. This test system may also be used to detect the specific effects on energy transduction, although this was not relevant to any compound investigated in this study. As examples of specific receptor-mediated toxicity, we chose the yeast estrogen screen (YES) as a specific test for estrogenicity, and the inhibition of chlorophyll fluorescence in algae to assess specific effects on photosynthesis. Reactive modes of action were assessed indirectly by measuring the relevance of cellular defense systems. Differences in growth inhibition curves between a mutant of Escherichia coli that could not synthesize glutathione and its parent strain indicate the relevance of conjugation with glutathione as a defense mechanism, which is an indirect indicator of protein damage. DNA damage was assessed by comparing the growth inhibition in a strain that lacks various DNA repair systems with that in its competent parent strain. Most compounds acted merely as baseline toxicants in all test systems. As expected, ethinylestradiol was the only compound showing estrogenic activity. Propranolol was baseline-toxic in all test systems exceptforthe photosynthesis inhibition assay, where it surprisingly showed a 100-fold excess toxicity over the predicted baseline effect. The exact mode of toxic action could not be confirmed, but additional chlorophyll fluorescence induction experiments excluded the possibility of direct interference with photosynthesis through photosystem II inhibition. Mixture experiments were performed as a diagnostic tool to analyze the mode of toxic action. Compounds with the same mode of toxic action showed the expected concentration addition. In the photosynthesis inhibition assay, agreement between experimental results and prediction was best for two-stage predictions considering the assigned modes of action. In a two-stage prediction, concentration addition was used as a model to predict the mixture effect of the baseline toxicants followed by their independent action as a single component combined with the specifically acting compound propranolol and the reference compound diuron. A comparison with acute toxicity data for algae, daphnia, and ...

Life cycle responses of the midge Chironomus riparius to polycyclic aromatic compound exposure

“…Several studies have shown that narcosis is strongly related to the lipophilicity of the compound, commonly expressed as the n-octanol-water partition coefficient (K ow ) (De Voogt et al, 1988). Consequently, deviations from the relationship between effect concentrations and K ow values may indicate a different mode of action (Bearden and Schultz, 1998;Escher et al, 2005). This was clearly demonstrated previously for soil invertebrates chronically exposed to several PACs (Sverdrup et al, 2001;Droge et al, 2006), and therefore in the present study we followed the same approach.…”

Bridging levels of pharmaceuticals in river water with biological community structure in the llobregat river basin (northeast spain)

“…Pharmaceuticals do not appear as isolated compounds in river water but as a mixture, and data on the responses of aquatic organisms to a mixture of pharmaceuticals are very limited. Escher et al [36] reported higher toxicity of a pharmaceutical mixture than separated products. Flaherty et al [37] indicated unpredictable and complex effects of a pharmaceutical mixture on Daphnia survival, growth, and reproduction.…”

Mode of Action Assignment of Chemicals Using Toxicogenomics: A Case Study with Oxidative Uncouplers

“…The question is how to recognize that two chemicals have a common MoA, and therefore can be grouped together? One possibility is to assign MoA based on molecular interactions between the chemical and the biological system which initiate specific toxicity pathways like binding to the estrogen receptors, or measuring specific cellular reactions that are indicative for specific effects such as chlorophyll interference in tests with algae (Hamadeh et al, 2002;Escher et al, 2005;Nendza and Wenzel, 2006;Woods et al, 2007;Pereira-Fernandes et al, 2014). Another approach is the grouping of chemicals on the basis of structural rules (Schüürmann, 1998;Verhaar et al, 2000;Enoch et al, 2008;Blackburn et al, 2011).…”