Electrochemical behavior of immobilized hemoglobin in alkaline solution

Abstract Glassy carbon electrode was modified with different synthesized hybrid clay-based materials and tested in alkaline solution with and without H 2 O 2 . The hybrid materials were obtained by immobilizing hemoglobin (Hb) on acid activated (AA) clay, or on AA clay modified with different sodium dodecyl sulfate (SDS) loadings. The obtained materials were characterized using DR UV–vis and ESR spectroscopy, elemental analysis, and SEM. The characterization confirmed higher degree of hemoglobin incorporation in the presence of SDS. The presence of SDS on the surface of clay particles resulted in the partial oxidation/denaturation of hemoglobin and formation of hemichrome. Cyclic voltammetry was used for the investigation of the electrochemical behavior of immobilized hemoglobin in alkaline solution. Two cathodic peaks at −0.45 V and −0.70 V were recorded and ascribed to the reduction of heme Fe(III)/Fe(II), and formation of HbFe(I) – highly reduced form of hemoglobin − respectively. The latter peak reflects hemoglobin denaturation. The presence of H 2 O 2 in the alkaline solution increased current intensities corresponding to both peaks (−0.45 V and −0.7 V). Linear response of peak current intensity vs. H 2 O 2 concentration was monitored for all investigated samples within different H 2 O 2 concentration ranges. The AA-SDS1.0-Hb electrode exhibited the highest current response with linear regression equation in the following form: I(μA) = 7.99 + 1.056 × [H 2 O 2 ] (mM) (R = 0.996). The limit of detection of 28 μM was estimated using the 3 sigma method. Different modified electrodes exhibited different degrees of denaturation resistance. The obtained values of Michaelis-Menten constant indicated that prolonged cycling in the presence of SDS increases protein denaturation.