Characterization of Oxidative Stress in Chicken Heart Tissue by High-Performance Liquid Chromatography with Ultraviolet‐Visible (HPLC–UV–Vis) Detection

Abstract

Oxidative stress can lead to tissue damage in the body, resulting in conditions that may lead to heart failure. Carbonyl proteins and malondialdehyde are common markers for determining oxidative stress. In this study, a high-performance liquid chromatography with ultraviolet‐visible (HPLC–UV–Vis) method was developed for both carbonyl proteins and malondialdehyde and compared with common spectrophotometric methods. The HPLC–UV–Vis method for determining carbonyl proteins showed a linear correlation in a range of 0.05–1.6 µg.mL−1 (correlation coefficient: 0.9939). The limits of detection and quantitation were determined to be 1.7 and 5.1 pmol on-column, respectively. The HPLC–UV–Vis method for malondialdehyde exhibited a linear correlation in the range of 0.04–13.6 µg.mL−1 (correlation coefficient: 0.9982) with limits of detection and quantitation of 1.5 and 4.5 pmol on-column, respectively. As a proof of concept, the applicability of the methods for both marker substances was investigated in chicken heart tissue as a complex matrix sample. To simulate oxidative stress, the tissue was treated with hydrogen peroxide as the oxidizing agent. Samples prepared with hydrogen peroxide showed increased carbonyl protein and malondialdehyde levels when compared to non-oxidized samples and samples treated additionally with the antioxidant Trolox. Regarding oxidative stress, similar results were achieved when compared with common spectrophotometric methods. However, HPLC–UV–Vis presented higher reproducibility and accuracy. The developed methods for both markers enable rapid and simple investigation of possible antioxidant effects on tissue samples.