Distinctively Different Effects of Perfluorobutanoic Acid and Perfluorononanoic Acid on Zebrafish Sex Differentiation and Androgen Receptor Activity

With the prohibition of perfluorooctanoic acid (PFOA), the emergence of alternative perfluoroalkyl substances such as perfluorobutanonic acid (PFBA) and perfluorononanoic acid (PFNA) in various environmental matrices has led to concerns about their adverse effects on humans and biota. This study aims to investigate the reproductive and developmental toxicity of PFBA and PFNA by combined <i>in vivo</i>, <i>in vitro</i>, and <i>in silico</i> approaches. Examination of juvenile zebrafish exposed to PFBA at environmental concentrations by histopathology, sex hormone, and gene expression revealed accelerated development of zebrafish toward males, while exposure to PFNA during sex differentiation resulted in feminization. In accordance with the <i>in vivo</i> results, PFBA activated the androgen receptor (AR) signaling pathway, but PFNA inhibited it in both prostate cancer cell proliferation and luciferase reporter gene assays. Similarly, the differential binding mode of the two chemicals to AR was shown in the molecular docking analysis, with PFBA exhibiting higher potency for the agonist conformation and PFNA favoring the antagonistic conformation. Together, these results suggest that, while PFNA exhibited similar effects on sex differentiation and AR activity as PFOA, PFBA showed distinctive effects and deserves particular attention and further investigation.