Feeding experiments were performed to investigate mercury (Hg) isotope fractionation during trophic transfer and internal distribution of total Hg (THg) in marine fish on exposure to natural seafood. Young-of-the-year amberjack (Seriola dumerili) were fed with either blackfin tuna (Thunnus atlanticus; 2647 ng/g THg) or brown shrimp (Farfantepenaeus aztecus; 25.1 ng/g THg) for 80 d or 50 d, respectively, and dissected for muscle, liver, kidney, brain, and blood. After 30 d of tuna consumption, Hg isotopes (δ(202) Hg and Δ(199)Hg) of the amberjack organs shifted to the tuna value (δ(202)Hg = 0.55‰, Δ(199)Hg = 1.54‰,), demonstrating the absence of Hg isotope fractionation. When amberjack were fed a shrimp diet, there was an initial mixing of the amberjack organs toward the shrimp value (δ(202)Hg = -0.48‰, Δ(199)Hg = 0.32‰), followed by a cessation of further shifts in Δ(199)Hg and a small shift in δ(202)Hg. The failure of Δ(199)Hg to reach the shrimp value can be attributed to a reduction in Hg bioaccumulation from shrimp resulting from feeding inhibition and the δ(202)Hg shift can be attributed to a small internal fractionation during excretion. Given that the feeding rate and Hg concentration of the diet can influence internal Hg isotope distribution, these parameters must be considered in biosentinel fish studies.