Neurodevelopmental Toxicity of Tris(2,4-di- <i>tert</i> -Butylphenyl)phosphate: Mechanistic Insights from Laboratory Models and Implications for Human Health

Abstract Organophosphite antioxidants (OPAs) and their transformation products, novel organophosphate esters (NOPEs), are ubiquitous environmental contaminants, yet their toxicological profiles remain poorly understood. This study systematically characterized the neurodevelopmental toxicity of these compounds and evaluated their associated environmental risks within the adverse outcome pathway framework. Molecular docking and radioligand binding assays identified integrin αvβ3 as a primary molecular target for all tested OPAs and NOPEs, with tris(2,4-di-tert-butylphenyl)phosphate (AO168═O) exhibiting high binding affinity. Exposure to AO168═O at environmentally relevant concentrations (≥50 μg/L) triggered significant behavioral deficits in zebrafish larvae, including reduced locomotion, altered exploratory preference, and irregular locomotor patterning. In vivo imaging and molecular analyses revealed that AO168═O induced motor neuron abnormalities and suppressed neurogenesis markers. Transcriptomic profiling elucidated that AO168═O disrupted MAPK and calcium signaling pathways via integrin αvβ3 targeting, driving the observed neurotoxicity. Crucially, the derived integrin αvβ3 binding threshold (0.7 μg/L) is comparable to reported environmental concentrations, providing a critical early warning for aquatic ecosystems. Furthermore, “Genes-to-Pathways” analysis demonstrated high conservation of this toxicological pathway between zebrafish and humans, highlighting a potential human health risk. These findings deliver strong evidence for the hazard assessment of AO168═O and offer valuable insights for managing other emerging NOPEs.