Pharmaceuticals in the Environment: Assessing Potential Risks to Fish

The widespread presence of human pharmaceuticals in aquatic ecosystems potentially poses a significant threat to vulnerable organisms, such as fish. To address this issue, the environmental risk assessment (ERA) of active pharmaceutical ingredients (APIs) is now integrated into various regulatory frameworks worldwide. However, the ERA process is often resource-intensive and furthermore raises ethical concerns over the use of experimental animals. Moreover, its effective implementation is hampered by limited understanding of organism-specific pharmacokinetics, including the absorption, distribution, metabolism and excretion of drugs. As a result, New Approach (non-animal) Methodologies (NAMs) are increasingly being explored and adopted. While these methodologies hold promise for addressing vital knowledge gaps, ongoing refinement and validation are essential to keep pace with technological advances and ensure robust protection of organismal and environmental health. This thesis focuses on assessing fish as sensitive receptors of pharmaceutical pollutants in the aquatic environment, identifying key factors that influence organismal susceptibility, and evaluating a series of NAMs for their potential to enhance understanding of fish-specific pharmacokinetics and support a more targeted, refined ERA process. The first phase of this work entailed a critical review of existing literature, which highlighted the fundamental role of species-specific pharmacokinetics in determining the susceptibility of fish to pharmaceutical exposure and effects. With metabolism being a key contributor in this regard, an in vitro rainbow trout (Oncorhynchus mykiss) primary hepatocyte monolayer culture system was applied to assess the hepatic clearance of five APIs with diverse physicochemistries and drug targets, namely propranolol (beta-blocker), quetiapine (antipsychotic), mycophenolic acid (MPA; immunosuppressant), clozapine (antipsychotic) and olaparib (antineoplastic). Inter-relationships between API exposure and drug clearance processes in trout hepatocytes were also investigated by quantifying selected gene transcripts and the activity of cytochrome P4501A – an important phase I drug-metabolising enzyme. In addition to affirming the hepatocyte monolayer culture system’s usefulness, the findings from this work underscore the importance of acknowledging organism-specific clearance capacities and pathways. The monolayer model was also critically evaluated against other in vitro liver models used to measure pharmaceutical clearance in fish, including subcellular fractions, hepatocyte suspensions and spheroids. This analysis demonstrated the strong utility and broad applicability of the various in vitro models, offering a complementary toolkit for refining pharmaceutical ERA and advancing understanding of fish hepatic physiology. In the next phase of work in this thesis, rainbow trout tissue samples were used to study tissue-specific bioconcentration of APIs, as well as changes in gene transcript levels and metabolic enzyme activity following in vivo exposures to three of the case-study APIs (quetiapine, MPA and olaparib). These studies revealed the complexity and heterogeneity of biological responses in whole organisms to chemical exposure and identified some distinct API clearance mechanisms between fish and humans. Measured API tissue concentrations from the in vivo studies were subsequently used to evaluate the predictive accuracy of a refined, generalised fish physiologically based kinetic (PBK) model. After incorporating species-specific physiological pH and in vitro-derived plasma protein binding data, however, the refined model demonstrated only marginal improvement over the original version. Consequently, further optimisation and validation of the model are warranted. Taken together, and pending further optimisation, the tools explored in this thesis show promise as effective and ethically sound approaches for generating data to support pharmaceutical ERA and broader eco(toxico)logical research. In these contexts, their potential applications include prioritising APIs for further testing and improving understanding of species-specific risk, thereby reinforcing the growing relevance of NAMs.<p></p>

Author dc:creator

• Chrisna Matthee (21052481)

• ecotoxicology
• ADME
• environmental risk assessment
• pharmaceuticals
• 3Rs
• non-animal methods

Statement dc:rights

• All rights reserved
• Open Access after 2027-09-19

Identifier

10779/exe.31812142.v1

OAI identifier oai:identifier

oai:figshare.com:article/31812142