SENSITIVITY AND SELECTIVITY OF THE ELECTROCHEMICAL DETECTION OF THE COPPER(II) COMPLEXES OF BIOACTIVE PEPTIDES, AND COMPARISON TO MODEL STUDIES BY ROTATING RING-DISC ELECTRODE

Post-column reaction of peptides with Cu(II) can be used for the elect rochemical detection of peptides as their biuret complexes. Understand ing of the behavior (sensitivity at the anode and cathode in the dual- series erectrochemical detector) of the system is facilitated through the observation of the rotating ring disc voltammetry of some model co mpounds. In operation, the anodic signal from the oxidation of the Cu( II)-peptide to the Cu(III) form can be used to detect peptides, or the downstream cathode can be used to detect the Cu(III) form. The signal s appear at about 0.4 V (anode) for tetra- and longer peptides, 0.65 V for tripeptides. The anode signal is augmented by tyrosine (oxidation at 0.4-0.5 V) and tryptophan (0.5-0.6 V). If the cathode is used as t he detector in a two working electrode cell, the sensitivity depends o n the stability of the Cu(III) product. This is peptide dependent, but the signal is significant and useful analytically. Twenty-three bioac tive peptides in two groups, naturally electrochemically active and na turally electrochemically silent, and several model compounds have bee n studied. Both naturally electrochemically active peptides (contain t yrosine and/or tryptophan) and naturally electrochemically silent pept ides have been studied. Chromatography with an acetonitrile gradient h as been used to separate the peptides in each group. Detection limits are for non-electroactive peptides in the range of 16-100 fmol (10- mu l injection 1.6-10 nM, 100 mu l injection 0.16-1.0 nM), and for elect roactive peptides in the range of 6-40 fmol (0.6-4.0 nM for a 10- mu l injection and 60-400 pM for a 100- mu l injection). A tryptic digest of bovine cytochrome c is easily seen at 100 nM.