APPLICATION OF COVALENT AFFINITY-CHROMATOGRAPHY WITH THIOL-DISULFIDE INTERCHANGE FOR DETERMINATION OF ENVIRONMENTAL EXPOSITION TO HEAVY-METALS BASED ON THE QUANTITATIVE-DETERMINATION OF ZN-THIONEIN FROM PHYSIOLOGICAL HUMAN FLUIDS BY INDIRECT METHOD BASED ON ANALYSIS OF METAL CONTENTS

Intoxication with heavy metals results in numerous poisonings and dise ases. They disturb metabolism of the system, are the source of cancer, degeneration changes and others. As a result of kidney damage the uri ne of people exposed to heavy metals contains different low molecular weight proteins, oligopeptides and amino acids, indicating pathologica l changes. One of the proteins is a very specific metallopolythiopolyp eptide-metallothionein (MT). Based on earlier investigations, a very g ood correlations has been found between the contents of metallothionei n in urine and plasma and the concentration of heavy metals in the blo od, urine, kidneys, liver and brain and general in level of exposition to heavy metals. The aim of our investigations was to carry out quant itative isolation of Zn-thionein (Zn-Th), in order to determine the le vel of exposition to heavy metals. For Zn-Th protein isolation by cova lent affinity chromatography with thiol-disulphide interchange (CAC-TD I) was applied, which is a modern technique of separation of a high af finity, good repeatability and reproducibility, allowing specific isol ation of the thiol-proteins CAC-TDI gel was used as a solid-phase extr action (SPE) support for preconcentration of Zn-Th protein and Zn bond ed with Zn-Th from water, rine, plasma and breast milk samples. The in vestigations showed unfavourable effect of the support on separation o f thiol proteins and good correlation between the concentration of MTs protein added to water, plasma and urine and the concentration of pro tein indirectly determined via atomic absorption spectrometric (AAS) m ethod, by preconcentration on SPE support metals formerly bound with M T protein and adsorbed on CAC-TDI gel and calculated from metals conce ntration. The present paper is a continuation of earlier experiments o n quantitation of Hg-thionein and Cd-thionein in physiological fluids and homogenates. (C) 1998 John Wiley & Sons, Ltd.