Some methodological aspects of metallothionein evaluation

The specificity of the methods used for metallothioneins (MTs) determination in both field and laboratory studies is crucial to a relevant interpretation of the results. The technique applied in the present study is commonly used, but several potential problems may limit its validity: (i) the stability of the metal-SH links during heating and the possibility of metal exchanges between MTs and others compounds; (ii) the presence of heat-stable thiol compounds with high molecular weight or very low molecular weight, which may subsist in the supernatant and interfere during global analysis of MTs; (iii) the subsistence of metals not bound to MT in the supernatant after heating; and (iv) the impact of reducing agents such as mercaptoethanol on the metal-binding ligands. Our investigations were conducted mainly on oysters (Crassostrea gigas) and polychaetes (Hediste/Nereis diversicolor) sampled, respectively, in two French metal-rich sites, the Gironde estuary and Boulogne harbour. This study, which allowed us to clarify most of the potential problems noted above, indicates the importance of dealing with methodological difficulties. (C) 2002 Elsevier Science Inc. All rights reserved.