HPLC-AAS HYBRID TECHNIQUE FOR STUDYING THE SPECIATION OF TRACE-METALS(AL, FE, SI, HG) IN BIOLOGICAL-FLUIDS - A REVIEW OF APPLICATIONS, RECENT EXPERIENCES AND PERSPECTIVES

A method developed using the combination of HPLC and AAS enabling both qualitative and quantitative study of the protein binding and speciat ion of trace metals in biological fluids at clinical relevant levels i s reviewed. The whole system was made metal-free by using polymer-base d columns, column holders and tubing and by the insertion of a silica- based scavenger column placed immediately before the injection valve t o selectively retain any trace of aluminum and iron originating from b uffer solutions and recipients. ETAAS instrumental conditions were car efully selected to eliminate interferences secondary to the salt gradi ent elution. Particular attention was paid to the choice of HPLC colum ns. Protein recoveries varied between 95 and 105% and trace metal reco veries were close to 100%. Intra-assay and inter-assay CVs of the HPLC /AAS hybrid technique were below 10%. Because of its high sensitivity, the method can be used at clinically relevant concentrations and was applied successfully to study (i) the interaction between iron and alu minum for binding to transferrin, (ii) the influence of citrate on the transferrin binding of aluminum, (iii) the speciation of silicon in t he serum of dialysis patients and (iv) the toxicity of mercury compoun ds in cell cultures.