Studies of potential-dependent metallothionein adsorptions using a low-volume electrochemical quartz crystal microbalance flow cell

Adsorptions of metallothioneins (MTs) onto thin mercury films (TMFs) at potentials more negative or positive than the point of zero charge (pzc) of mercury were quantified for the first time, using a novel electrochemical quartz crystal microbalance (EQCM) flow cell in conjunction with a simple flow-injection system (FI-EQCM). The EQCM flow cell has a low internal volume (ca. 50 μl) designed for the investigation of biomolecular attachment to surfaces. Such a cell facilitates in-situ gravimetric measurements of MT adsorption processes at different potentials. The amount of the MT adsorption at −0.9 V, a potential more negative than the pzc of the TMF, was found to be greater than that attached via simple chemisorption (in an open-circuit configuration). The quantity of metals released by the MT adsorbates, measured by electrochemical inductively coupled plasma-atomic emission spectrometry (EC/ICP-AES), was found to constitute ∼3.8% of the total mass measured by FI-EQCM. The average number of cysteines per MT molecule involved in the Hg-cysteine thiolate formation was found to be 4.2±1.8. Thus, it appears that about 1–2 metal ions (Cd2+) per MT molecule have been released off the TMF electrode since the stoichiometric ratio between the Cd2+ and the cysteines in a MT molecule is known to be 1/3. The quantitative measurements by the two hybrid electrochemical methods (FI-EQCM and EC/ICP-AES) help elucidate the electrode reactions of MT adsorbates at TMF surfaces and the MT metal transfer processes.