Evaluation of different purification procedures for the electrochemical quantification of mussel metallothioneins.

Several different procedures were applied to purify metallothionein (MT) isolated from the digestive glands of the mussels (Mytilus galloprovincialis) exposed to cadmium: heat treatment (at 70 and 85 degrees C), solvent precipitation, and gel-filtration. Using electrochemical method, the MT contents were determined. Based on the obtained results each procedure was statistically evaluated. Application of the post-hoc Scheffé test showed that there were significant differences between all the treatments applied. Heat treatment and solvent precipitation effectively remove high molecular weight (HMW) proteins from the samples. In untreated as well as in heat or solvent treated samples MT10 isoform remains unchanged. MT20 isoform is significantly reduced by heat treatment and drastically reduced by solvent precipitation. Due to the fact that MT20 is considered as 'target' MT isoform for metal effect and exposure it is recommended to use the purification procedure which less affects MT20 fraction. According to the presented results it is the heat treatment.