Tez özetleri Astronomi ve Uzay Bilimleri Anabilim Dalı

Application of CUPRAC spectrophotometric method to antioxidant compounds containing thiol groups

In the course of oxidative metabolism, oxygen may produce reactive oxygen species (ROS) such as singlet oxygen, superoxide anion and hydroxyl radicals in the human body. Antioxidants are health beneficial compounds (through their combat with ROS) that significantly inhibit or retard such adverse reactions, and therefore, determination of total antioxidant capacity is important. Thiol-type compounds (containing sulfur) are an important class of strong antioxidants. In this regard, the aim of this thesis work is to to apply the CUPRAC (cupric ion reducing antioxidant capacity) assay to thiol-type antioxidants and comparison of the antioxidant capacities with those found by the reference assays for method validation.

When the CUPRAC method was applied to these compounds, their antioxidant capacity and corresponding TEAC (trolox equivalent antioxidant capacity) values were determined. In this regard, the CUPRAC TEAC values of thiol-type antioxidants in descending order could be listed as: 1,4-dithioerythritol (DTE: 2,3-dihydroxybutane-1,4-dithiol), reduced glutathione (GSH), dihydrolipoic acid (DHLA), homocysteine, N-acetyl cysteine (NAC), cysteine, cysteamine, glutathione ethyl ester (GSHEE). Synthetic mixtures of these thiols were prepared, and the agreement between the expected and found TAC values (in mM trolox-TR equivalents) of these mixtures were compared, yielding coherent results. The technique of standard additions was performed for thiol mixtures, and the results confirmed that apparent chemical deviations from Beer’s law were absent in the system. The CUPRAC results were compared with those of reference methods, namely ABTS and FRAP. As being one of the most important thiol (-SH) peptide at ‘in vivo’ conditions, glutathione showed a TEAC value of 0.57. On the other hand, the ABTS TEAC value of GSH was 1.51. The ABTS result was not in accordance with the reversible 1-e reduction of GSH to the corresponding disulfide (2GSH ↔ GSSG + 2H⁺ + 2e^-) that is expected to occur under physiological conditions. FRAP did not give consistent results, and even at relatively high concentrations of GSH, the FRAP TEAC value was only 0.07.

CUPRAC and reverse phase HPLC (high performance liquid chromatography) methods were applied to thiol-type antioxidant−bearing pharmaceuticals, and the obtained results were compared. The pharmaceuticals of Brunac eye drop, Trom and Mentopin effervescence tablets containing NAC were assayed with HPLC for comparison, and the obtained results for NAC and AA (ascorbic acid) were in accordance with those found with CUPRAC.