Analysis of pyocyanin from Pseudomonas aeruginosa by adsorptive stripping voltammetry

Abstract

We have developed an adsorptive stripping voltammetric (AdSV) method that can be used for analysis of pyocyanin at the nanomolar level. The method employs a hanging mercury drop electrode (HMDE) as a working electrode in a three-electrode system in a 15-mL quartz cell. Preconcentration of pyocyanin in a 10-mM ammonia buffer (pH 8.0) on the HMDE is carried out by adsorption under conditions of controlled mass transfer for 60 sec, followed by scanning the electrode potential that results in reduction (cathodic scan) or oxidation (anodic scan) of the accumulated pyocyanin. For analysis of pyocyanin in samples containing a significant amount of surface-active species from a bacterial culture broth, a positive-going (anodic) potential scan must be applied after an adsorption at −0.50 V. Anodic scan of the adsorbed pyocyanin provided a well-defined oxidation peak at a potential of −0.17 V. The “anodic scan” variant of the method was tested for utility in Mueller-Hinton broth diluted from 20 to 200 times in a 10-mM ammonia buffer. Reproducibility of the method as applied to the analysis of pyocyanin produced by Pseudomonas aeruginosa (18.83 ± 0.32 μM) in culture was demonstrated by the within-day and day-to-day coefficients of variation of 1.7% and 3.4%, respectively.