Long-term exposure to 2,4-di-tert-butylphenol impairs zebrafish fecundity and affects offspring development

As a widely used antioxidant, 2,4-di-tert-butylphenol (2,4-DTBP) has been frequently detected in the environment and biota. Although a few studies reported its hormone-like activity in vitro, the endocrine disrupting potential of 2,4-DTBP and its effect on reproduction are not yet elucidated. In this study, adult zebrafish were exposed to 5 and 50 nM 2,4-DTBP for 60 days. Reduction in cumulative egg production was observed after 45 days of exposure. Gonadal maturation was also delayed in both female and male zebrafish following 2,4-DTBP exposure. The impaired fecundity was attributed to an imbalance of 17β-estradiol/testosterone ratio (E2/T) and altered transcripts involved in the hypothalamic-pituitary-gonadal (HPG) axis. Upon exposure, aromatase (CYP19) and E2 levels were significantly decreased in females, but were increased in males. Additionally, molecular docking revealed potential binding of 2,4-DTBP to estrogen receptors and CYP19, highlighting molecular initiating events that may interfere with steroid hormone synthesis. We also showed that 2,4-DTBP can be transferred to offspring, affecting their development and compromising immunity. The expression of triiodothyronine (T3) and hatching-related genes (esr2α, esr2β, and zhe2) were altered, suggesting that parental exposure to 2,4-DTBP resulted in intergenerational toxicity in F1 larvae. Taken together, these findings provide novel insight into the reproductive toxicity of 2,4-DTBP, contributing to its ecological risk assessment.