Enhancing effect of melatonin on chemiluminescence accompanying decomposition of hydrogen peroxide in the presence of copper

• Published in: Free radical biology & medicine Vol. 34; no. 12; pp. 1544 - 1554
• Main Authors: Kładna, Aleksandra, Aboul-Enein, Hassan Y., Kruk, Irena
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 15.06.2003
• Subjects: Chemiluminescence; Copper - metabolism; Electron Spin Resonance Spectroscopy; EPR spectroscopy; Fluorescence; Free Radical Scavengers - pharmacology; Free radicals; Hydrogen Peroxide - metabolism; Luminescent Measurements; Melatonin; Melatonin - pharmacology; Reactive Oxygen Species; Spectrophotometry; Spin Labels; Spin Trapping; Melatonin; Reactive oxygen species; Free radicals; Fluorescence; Chemiluminescence; Spectrophotometry; EPR spectroscopy
• ISSN: 0891-5849; 1873-4596
• DOI: 10.1016/S0891-5849(03)00180-1

The oxidation of melatonin (MEL) using the Cu(II) + H 2O 2 + HO − (the Fenton-like reaction) system was investigated by chemiluminescence (CL), fluorescence, spectrophotometric, and EPR spin trapping techniques. The reaction exhibits CL in the 400–730 nm region. The light emission from the Fenton-like reaction was greatly enhanced in the presence of MEL and was strongly dependent on its concentration. The spectrum measured with cut-off filters revealed maxima at around 460, 500, 580–590, 640–650, and 690–700 nm. The band at 460 nm may be due to the excited cleavage product, N 1-acetyl-N 2-formyl-5-methoxykynuramine, whereas the bands at 500, 580–590, 640–650, and 700 nm were similar to those observed for singlet molecular oxygen ( 1O 2). The effect of reactive oxygen species (ROS) scavengers on the light emission was studied. The CL was strongly inhibited by the 1O 2 scavengers in a dose-dependent manner; at concentration 1 mM the potency of 1O 2 scavenging was 5,5-dimethylcyclohexandione-1,3 > methionine > histidine > hydroquinone. The potency of HO • scavenging by thiourea, tryptophan, cysteine at concentration 5 mM was 79–94%, by 1 mM glutathione and trolox 75 and 94%, respectively, and by 10 mM cimetidine 18%. Specific acceptors of O 2 • − such as p-nitroblue tetrazolium chloride and 4,5-dihydroxy-1,3-benzene disulfonic acid (tiron) at concentration 5 mM decreased the CL by 51 and 95%, respectively, whereas superoxide dismutase (SOD) does not reduce the emission at concentration 2.8 U/ml. At higher concentration SOD substantially enhanced the light emission. Addition of 1360 U/ml catalase and 100 μM desferrioxamine strongly inhibited CL (96 and 90%, respectively). The increased generation of 1O 2 from the Cu/H 2O 2 system in the presence of MEL was confirmed using the spectrophotometric method based on the bleaching of p-nitrosodimethylaniline and by trapping experiments with 2,2,6,6-tetramethylpiperidine (TEMP) and subsequent electron paramagnetic (EPR) spectroscopy. These findings suggest the increased production of reactive oxygen species (O 2 • −, HO •, 1O 2) from the Fenton-like reaction in the presence of MEL. This means that the hormone is not able to act as classical chain-breaking antioxidant even at low concentration, and may show clear prooxidant activity at higher concentrations. In addition, long-lived carbonyl product of the MEL transformation in the triplet state can also be toxic by transferring its energy to organelles and causing a photochemical process.