Characterization of Hg(II) binding with different length phytochelatins using liquid chromatography and amperometric detection.

A simple and rapid methodology is optimised to analyse mixtures of different phytochelatins (PC(n), n=2-5) with Hg(II) by HPLC with amperometric detection as a first step towards the analysis of extracts of plants stressed with Hg(II). The separation was achieved in a C(18) column with a mobile phase of 0.1% trifluoroacetic acid (TFA) in water and 0.1% TFA in acetonitrile using gradient elution. Electrochemical detection with glassy carbon electrode and UV-vis detection were used in series. This methodology can clearly distinguish between the free peptides and their complexes and permits to study the evolution of the different complexes formed and predicts the possible interactions between the long chain phytochelatin complexes. ESI-MS is used as a complementary technique to find out the stoichiometries of such long chain phytochelatin complexes.