Electrochemical Sensor for Carbonyl Groups in Oxidized Proteins

Abstract

The interaction of proteins with free radicals leads, among other types of damages, to the production of stable carbonyl groups, which can be used as a quantification of oxidative stress at proteins level. The aim of this study was the development of an electrochemical sensor for the detection of carbonyl groups in proteins oxidized by reactive oxygen species. Its working principle is based on the redox properties of dinitrophenylhydrazine (DNPH). BSA was used as a model protein and its oxidation achieved through Fenton reactions. Using differential pulse voltammetry at glassy carbon electrode, the electrochemical behavior of DNPH and of the native and oxidized BSA was investigated in solution. It has been shown that the hydrazine moiety of the DNPH is the electroactive center and is responsible for carbonyl complexation. Special attention was paid to the immobilization of the DNPH in order to retain its redox properties, and this was achieved on a mixed 4-styrenesulfonic acid-nafion matrix. The sensor’s surface characterization and the detection of carbonyl groups in oxidized protein were performed by voltammetry, Fourier-transformed infrared spectroscopy and scanning electron microscopy while the voltammetric results were confirmed by surface plasmon resonance measurements. It has been shown that upon interaction with carbonyl groups of the oxidized protein, the oxidation peak of the hydrazine moiety of DNPH decreases as a function of incubation time and protein concentration. The sensor sensitivity was 0.015 nmol carbonyl per mg of oxidized protein and detection limits of 50 μg oxidized BSA and 0.75 pmol carbonyls.

Coaxial dual-path electrochemical biosensing and logic strategy-based detection of lung cancer-derived exosomal PD-L1

“…Compared with traditional detection methods such as fluorescence, the use of electrochemical biosensors has gained widespread attention in marker detection due to their high sensitivity and ease of operation, 6,7 but they still face challenges in the detection of low abundance ExoPD-L1. Advances in nanotechnology have enabled nanomaterials to show outstanding advantages in highly sensitive detection of electrochemical biosensors.…”

Electrochemical approaches based on micro- and nanomaterials for diagnosing oxidative stress

“…[5]. This pathological or physiological process generates certain electroactive molecules known as biomarkers via hydroxylation, nitration [6], oxidative deamination (reactive carbonyl moiety) [7], and free radical chain oxidation (O 2 addition to carbon radicals; fragmentation of peroxyl radical) [8] reactions. Oxidative stress can be diagnosed by monitoring oxidation adducts of DNA, proteins, lipids, the activity of enzymes, and antioxidants status [9,10] (Scheme 1).…”

Carbon Inks-Based Screen-Printed Electrodes for Qualitative Analysis of Amino Acids

“…Until the end of 1980s, it was already demonstrated that the redox properties of amino acids, especially Tyr and Trp, allow the investigation of proteins [ 1 , 2 ]. The last decade came with the elucidation of electroactive amino acids oxidation mechanisms [ 5 , 6 , 7 , 8 , 9 ], while the research was extended to broad studies regarding different phenomena such as adsorption [ 9 ], aggregation [ 10 ], fibrilization [ 11 ], structural modifications [ 12 ], protein/enzyme–substrate interaction [ 13 ], etc., and the investigation addressed to peptides [ 14 ], amyloids [ 15 , 16 ], proteins [ 15 , 17 ], and enzymes [ 14 ]. At the same time, in the last decade, this field of electrochemistry has been governed by the use of glassy carbon working electrodes [ 18 ] and other carbon electrodes, such as disposable carbon-based screen-printed electrodes [ 19 ], mainly due to their broad potential window, which is usual between −1.0 V and +1.4 V [ 18 , 20 , 21 , 22 ].…”