Influence of tyrosine on the dual electrode electrochemical detection of copper(II)-peptide complexes

Abstract

The bluret reaction makes peptides electrochemically active. This is the basis of an electrochemical method for the detection of peptides following their liquid chromatographic separation. This paper discusses the influence of tyrosine, an electroactive amino acid, on the detection of Cu(II)-peptide (bluret) complexes containing it. The dual electrode detector has an upstream anode and a downstream cathode. Tyrosine-containing peptides yield anodic signals that are approximately the sum of the tyrosine signal and the signal from the bluret complex of a nonelectroactive model peptide, phenylalanylglycyl-glycine (FGG). The cathodic signal is depressed in comparison to FGG. This is traced to the presence of an intramolecular reaction between Cu(III) and a reaction product resulting from the oxidation of the tyrosinyl residue. The rate constant for the corresponding intermolecular reaction is significant (10(6)-10(7) M-1 s-1), but in practical analytical situations, the concentrations of the reactants will be small, so the reaction will not be a major factor. Sensitivities for several bioactive peptides are reported. The dependence of the signals on the position of tyrosine in a tripeptide is also studied.

Separation and detection of angiotensin peptides by Cu(II) complexation and capillary electrophoresis with UV and electrochemical detection

“…The biuret reaction and LC-EC has been extensively investigated for the sensitive and selective determination of peptides [12][13][14][15][16][17][18][19][20]. The reaction conditions for complex formation had previously been investigated by both Margerum and co-workers [21][22][23][24] and Weber's group [25].…”

“…Most work using the biuret reaction has relied on preor postcolumn derivatization of peptides with copper to form copper-peptide complexes [12][13][14][15][18][19][20]. Recently, our group has shown that complexes could be formed on-capillary [28][29][30].…”

Detection of neuropeptides using on-capillary copper complexation and capillary electrophoresis with electrochemical detection

“…4). The Tyr-containing peptide copper complex provides a considerably greater electrochemical response than does the non Tyr-containing peptide copper complex, as expected [13,25]. However, the significant difference in E max illustrates the possibility for tunable selectivity with electrochemical detection.…”

“…The increase in sensitivity and lower LOD are due to a combination of factors. The anodic signal generated from the oxidation of the copper complex is an additive effect from the Tyr and Cu(II) oxidation [13]. The position of Tyr in the peptide also affects the electrochemical signal.…”

“…The position of Tyr in the peptide also affects the electrochemical signal. Peptides, such as Leu-enkephalin, containing a Tyr at the amine terminus have higher sensitivities than peptides with Tyr in nonterminal positions [13]. On-capillary complexation provides the added benefit of peak stacking [20].…”