Assessment of Early-Stage Fluorene Toxicity in Zebrafish Embryos Using an IrO₂/GCE-Based Electrochemical Sensor

Zebrafish (Danio rerio) is a widely adopted vertebrate model in toxicological research, yet current growth-toxicity assessments rely heavily on morphological endpoints that demand long observation periods and thus preclude rapid screening or early warning. The persistent organic pollutant fluorene perturbs purine metabolism in zebrafish embryos at environmentally relevant concentrations and subsequently impairs embryonic development. Herein, we present a novel electrochemical sensor based on an iridium oxide-modified glassy carbon electrode (IrO₂/GCE) that enables ultrasensitive monitoring of xanthine and guanine (X/G) fluctuations as metabolic biomarkers of fluorene cytotoxicity. Comprehensive characterization by scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDS), X-ray photoelectron spectroscopy (XPS), and cyclic voltammetry (CV) confirmed the surface morphology, elemental composition, and electrochemical performance of the modified electrode, affording high sensitivity and a wide linear range for the simultaneous detection of four purines. Using X/G as sentinel analytes, the IrO₂/GCE platform successfully tracked dynamic purine metabolic changes in fluorene-exposed embryos, corroborated by high-performance liquid chromatography (HPLC). Notably, the electrochemical assay already detected fluorene-induced suppression of purine levels at as early as 24 hpf, with excellent concordance to HPLC data. The IrO₂/GCE sensing system therefore provides a rapid, reliable, and embryo-scale tool for developmental toxicity assessment of fluorene and lays a solid technical foundation for the early-warning monitoring of persistent organic pollutants in the environment.