Voltammetric Determination of Leucoindigo Adsorbed on Pretreated Carbon Paste Electrodes: Its Application in a Flow System

Abstract

The determination of leucoindigo adsorbed on the surface of pretreated carbon paste electrodes has been performed by cyclic (CV) and alternating current voltammetry (ACV). The water-soluble compound called leucoindigo is obtained through the reduction of water insoluble indigo in an alkaline media and in presence of a dithionite salt (Na2S2O4) as a reducing agent. Cyclic voltammograms of leucoindigo show two reversible electrodic processes, in the aqueous 0.1 M Tris-HCl pH 7.2, at the formal potential of −0.4 V and +0.3 V (vs. Ag/AgCl/sat. KCl). In a batch protocol, leucoindigo was quantified by CV with a change of medium between accumulation and detection steps. Reversible voltammetric processes of leucoindigo were greatly enhanced by other voltammetric technique like alternating current voltammetry. Limit of detection in the nanomolar range was achieved for a 2 min accumulation time by ACV in a batch procedure. A flow system was also employed, with adsorptive voltammetric detection of leucoindigo, since this automates the methodology and decreases analysis time. Parameters related to the electrochemical technique are optimized and calibration plots obtained are reported. These data provide useful information about the suitability of using leucoindigo in the detection system of alkaline phosphatase (AP) and horseradish peroxidase (HRP) based affinity devices, in which indigo is generated by the enzymatic hydrolysis of the 3-indoxyl phosphate substrate.