Mercury isotope dynamics in marine fish during bioaccumulation and excretion: Temporal responses across tissues

Mercury (Hg) is a potent neurotoxin that bioaccumulates through aquatic food webs. In fish, Hg excretion regulates Hg burden, yet whether it induces tissue-specific Hg isotope fractionation remains unclear. To address this question, we conducted a controlled feeding experiment exposing starry flounders (Platichthys stellatus) to diets amended with inorganic mercury (IHg; 4 and 25 μg/g) or methylmercury (MeHg; 1 μg/g) for 4 weeks (bioaccumulation), followed by 16 weeks of Hg-unamended feeding (excretion). Total Hg and MeHg concentrations, together with Hg isotope ratios (δ²⁰²Hg, Δ¹⁹⁹Hg) were measured over time in the muscle, intestine, liver, kidney and feces. During bioaccumulation, near-complete Δ¹⁹⁹Hg equilibration with the respective diets was observed in all tissues except the muscle of IHg-fed fish, whereas δ²⁰²Hg exhibited treatment-specific fractionation. Under MeHg treatment, hepatic demethylation and subsequent redistribution of newly formed IHg caused mass-dependent fractionation (MDF) in the liver, kidney, and feces. Under IHg exposure, MDF was associated with glutathione-Hg binding and subsequent tissue redistribution, and higher IHg exposure further indicated intestinal methylation, as evidenced by negative δ²⁰²Hg shifts (-0.50 ‰) and elevated intestinal %MeHg (65%) despite the absence of external MeHg input. During excretion, the intestine and feces showed the most rapid Δ¹⁹⁹Hg turnover, whereas the muscle, liver, and kidney exhibited slower isotope turnover following the dietary shift. Under high IHg exposure, minor Δ¹⁹⁹Hg changes and distinct serial δ²⁰²Hg shifts across tissues suggest two-phase elimination, with initial loss of labile MeHg followed by removal of recalcitrant IHg, delaying Hg turnover across tissues, including the liver and kidney. Together, these results show that Hg excretion induces tissue-specific Hg isotope fractionation and turnover in fish, with patterns governed by Hg exposure level, speciation, and internal processing, providing toxicological insights into Hg persistence and fish tissue selection for biomonitoring.