Typical Thermochromic Dye Crystal Violet Lactone Acts as an Endocrine Disruptor: Nontargeted Screening, Prioritization, and <i>In Vivo</i> Experimental Verification

Identification and characterization of endocrine disruptors and their associated ecological and health risks are critical areas of environmental toxicology. In this study, we aimed to identify previously unrecognized endocrine disruptors in thermal paper by employing a nontargeted screening coupled ToxPi Ranking (NSTR) approach. Among the substances identified with high confidence in thermal paper, Crystal Violet Lactone (CVL), a typical thermochromic dye, was prioritized with a high endocrine disruptive potential. CVL can be absorbed through the skin and excreted in the urine. In addition, we demonstrated significant bioaccumulation of CVL in zebrafish tissues, with hepatic concentrations reaching up to 3.1 times those in the exposure water, and five biotransformation products were identified. CVL exposure impaired the reproductive capacity of breeding pairs, led to gonadal histological alterations, and exhibited developmental toxicity to F1 offspring. Transcriptional analysis revealed disruptions in the expression of hypothalamic-pituitary-gonad-liver (HPG-L) axis genes in adults. In embryos, CVL significantly increased the frequency of epiboly, yolk extension formation, and somite number while decreasing the ear-eye distance and size of the yolk sac. Significant alterations in HPG-L axis genes were observed, as well. Thus, our data demonstrated that CVL targeted multiple end points in both adult fish and embryos, exhibiting remarkably endocrine disruptive effects.