Stereoselective disruption of copper homeostasis by prothioconazole induces cardiotoxicity via mitochondria-mediated apoptosis in zebrafish embryos.

Chiral pesticides may exhibit enantioselectivity in biological activity, environmental fate, and toxicity. In this study, the enantioselective cardiotoxicity and underlying mechanisms of the chiral fungicide prothioconazole (PTCZ) were thoroughly investigated at the molecular level using zebrafish embryos. Results demonstrated significant stereoselectivity, with S-(+)-PTCZ inducing severe cardiac developmental toxicity, while R-(-)-PTCZ exhibited effects comparable to the control. Mechanistic studies revealed that S-(+)-PTCZ triggered profound copper homeostasis disruption, evidenced by decreased copper ions and downregulated expression of the copper transporter CTR1. This copper deficiency led to suppression of key copper-dependent enzymes, resulting in mitochondrial respiratory chain dysfunction. Consequently, mitochondrial membrane potential collapsed, mitochondrial quantity decreased, ATP production plummeted, and tricarboxylic acid (TCA) cycle intermediates were reduced. This cascade ultimately precipitated mitochondrial damage, cytochrome c release, an increased Bax/Bcl-2 ratio, activation of cleaved caspase-3, and cardiomyocyte apoptosis. In contrast, R-(-)-PTCZ exposure caused minimal disturbance to these pathways. This study demonstrates the stereoselective cardiotoxicity of S-(+)-PTCZ in fish and identifies copper homeostasis disruption as the key initiating event leading to energy crisis and apoptosis. These findings are crucial for accurate enantiomer-specific risk assessment of chiral pesticides and the development of safer, high-efficacy enantiopure agrochemicals.