Effect-based spatiotemporal assessment of suspended particulate matter in the River Rhine: An early warning platform for environmental monitoring

Effective early warning systems for aquatic contamination require monitoring strategies capable of detecting subtle, long-term shifts in mixture-driven biological activity. Suspended particulate matter (SPM) serves as a carrier and reservoir for complex contaminant mixtures, facilitating their transport and persistence in aquatic systems, yet systematic toxicological time series for archived SPM remain scarce. Regulatory monitoring predominantly targets Priority Substances and River Basin Specific Pollutants, leaving the temporal trends of particle-associated mixture toxicity largely unresolved. Leveraging 18 years (2005–2022) of cryogenically archived annual SPM composites from the Rhine River, we conducted a spatiotemporal effect-based assessment integrating receptor-mediated effects, oxidative stress analysis and untargeted Cell Painting phenomics. This integrated toolbox enabled evaluation of pathway-specific responses and multi-compartment cellular perturbations associated with particle-bound contaminant mixtures. Polar SPM-associated chemicals elicited oxidative stress response and caused endocrine disruption through estrogen receptor α (ERα) activation and androgen receptor inhibition (anti-AR). Trend analysis showed spatiotemporal variation along the river, with statistically increasing trends of oxidative stress and anti-AR activity over time at Koblenz, driven by polar chemicals. Both polar and non-polar SPM extracts activated the aryl hydrocarbon receptor (AhR), indicating presence of compounds capable of triggering xenobiotic response pathways. Several subcellular compartments were affected, with mitochondrial features being among the most affected. These findings demonstrate that SPM-associated chemicals elicit diverse toxicological effects by acting on several receptors and impacting diverse cellular structures. Combining targeted and phenomics-based effect approaches provided comprehensive mechanistic insights and valuable information to support the early warning systems for chemical contamination in aquatic environments.