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The group of per-and polyfluoroalkyl substances (PFAS) comprises thousands of chemicals, which are used in various industrial applications and consumer products. In this study, a feeding experiment with laying hens and feed grown on a contamination site was conducted, and PFAS were analyzed in the feed and eggs to assess the transfer of PFAS into eggs. A targeted analysis of perfluoroalkyl acids (PFAAs) and different sulfonamides was performed. Additionally, the total oxidizable precursor (TOP) assay was modified by fully oxidizing small amounts of the samples instead of oxidizing their extracts in order to overcome potential losses during extraction. Targeted analysis showed the presence of known PFAAs and four sulfonamides in the feed and egg yolk samples. In the plant-based feed, short-chain PFAAs, methyl and ethyl perfluorooctane sulfonamidoacetic acid (Me-and EtFOSAA), and perfluorooctane sulfonic acid (PFOS) were the most abundant PFAS. In the eggs, PFOS, FOSAA, and its alkylated homologues showed the highest concentrations. The TOP assay revealed the presence of substantial amounts of precursors with different chain lengths from C4 to C8. The highest relative increase of PFOA after oxidation was observed in egg yolk from the end of the exposure period (828%). The results of this study demonstrate the transfer of PFAAs and their precursors into hens' eggs and emphasize the contribution of (known and unidentified) precursors to the overall PFAS burden in edible products. The modified TOP assay approach was shown to be a powerful tool to better assess the total burden of samples with PFAS.

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