The sediments from Chinese coastal waters contain relatively high concentrations of arsenic (As), mainly arsenate As(V), which may be transferred along the marine benthic food chain. The prey-specific determination of As bioaccumulation and transformation in marine benthic fish remains little known. In this study, we focused on a typical marine benthic food chain comprising of sediments, deposit-feeding invertebrates (polychaete Nereis succinea and clam Gafrarium tumidum) and goby fish Mugilogobius chulae. Graded exposed experiments using different As exposure durations and concentrations were conducted to examine their transformation rate and efficiency. Radiotracer techniques were used to determine the rates of As uptake (as arsenate) from seawater, assimilation from two prey and its subsequent efflux in the goby fish. We demonstrated that the two prey (polychates and clams) displayed different As biotransformation in the goby fish. Biotransformation rate was higher in the goby fish fed on the clams than on the polychaetes, and biotransformation efficiency was lower with increasing inorganic As concentration in the prey. The As overall bioaccumulation in the goby fish was very low, mainly because of the low dissolved uptake and dietary assimilation and high efflux. Combining the biotransformation and biokinetics measurements, our findings highlighted that different prey containing different As concentrations and As species resulted in the comparable As bioaccumulation in the goby fish.