Trace metal speciation in seawater from the Bulgarian Black Sea coast was studied in situ by hollow fiber permeation liquid membrane (HF-PLM) and by diffusion gradients in thin-film gels (DGT). The concentrations of Cd, Cu, Ni, and Pb determined by HF-PLM were lower than those measured by DGT, in agreement with their analytical windows, e.g., free metal ions provided by the HF-PLM and dynamic (mobile and labile) species by the DGT. The obtained suite of data was further used to evaluate the bioavailability of these metals to the microorganisms, which was then compared with experimental results of metal uptake to green microalga Chlorella salina. Uptake fluxes of the Cd, Cu, Ni, and Pb to C. salina,were predicted from the measured HF-PLM concentrations and laboratory experimentation in artificial seawater, in agreement with theoretical considerations. The HF-PLM and DGT appear to be promising analytical techniques for speciation and bioavailability studies in complex environmental media and allow improved understanding of the role of different chemical species in metal bioavailability (and impact) in seawaters.