Reaction mechanism of melatonin oxidation by reactive oxygen species in vitro.
ABSTRACT Melatonin (N-acetyl-5-hydroxytryptamine) is a pineal hormone widely known for its antioxidant properties, both in vivo and by direct capture of free radicals in vitro. Although some metabolites and oxidation products of melatonin have been identified, the molecular mechanism by which melatonin exerts its antioxidant properties has not been totally unravelled. This study investigated the reaction mechanism of oxidation of melatonin by radio-induced reactive oxygen species, generated by gamma radiolysis of water for aqueous solutions of melatonin (from 20 to 200 μm), in the presence or absence of molecular oxygen. The hydroxyl radical was found to be the unique species able to initiate the oxidation process, leading to three main products, e.g. N(1)-acetyl-N(2)-formyl-5-methoxykynurenin (AFMK), N(1)-acetyl-5-methoxykynurenin (AMK) and hydroxymelatonin (HO-MLT). The generation of AFMK and HO-MLT strongly depended on the presence of molecular oxygen in solution: AFMK was the major product in aerated solutions (84%), whereas HO-MLT was favoured in the absence of oxygen (86%). Concentrations of AMK remained quite low, and AMK was proposed to result from a chemical hydrolysis of AFMK in solution. A K-value of 1.1 × 10(-4) was calculated for this equilibrium. Both hydrogen peroxide and superoxide dismutase had no effect on the radio-induced oxidation of melatonin, in good accordance for the second case with the poor reactivity of the superoxide anion towards melatonin. Finally, a reaction mechanism was proposed for the oxidation of melatonin in vitro.
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ABSTRACT: This study was performed to investigate the effect of melatonin on bovine oocyte maturation and subsequent embryonic development in vitro. The endogenous melatonin concentration in bovine follicular fluid is approximately 10-11 M. To examine the potential beneficial effects of melatonin on bovine oocyte maturation in vitro, germinal vesicle (GV) oocytes was incubated with different concentrations of melatonin (10-11, 10-9, 10-7, 10-5, 10-3M). Melatonin supplementation at suitable concentrations significantly promoted oocyte maturation. The development of embryos and the mean cell number/blastocyst produced after in vitro fertilization were remarkablely improved. The most effective melatonin concentrations obtained from the studies ranged from 10-9 to 10-7 M. The expression of melatonin receptor MT1 and MT2 gene was identified in cumulus cells, granulosa cells and oocytes using reverse transcription PCR, immunofluorescence and western blot. The mechanistic studies show that the beneficial effects of melatonin on bovine oocyte maturation are mediated via melatonin membrane receptors since the melatonin receptor agonist (IIK7) promotes this effect while the melatonin receptor antagonist (luzindole) blocks this action. Mechanistic explorations revealed that melatonin supplementation during bovine oocyte maturation significantly up-regulated the expressions of oocyte maturation associated genes (GDF9, MARF1 and DNMT1a) and cumulus cells expansion related genes (PTX3, HAS1/2) and that LHR1/2, EGFR are involved in signal transduction and epigenetic reprogramming. The results obtained from the studies provide new information regarding the mechanisms by which melatonin promotes bovine oocyte maturation in vitro, and provide an important reference for in vitro embryo production of bovine and the human assisted reproductive technology.This article is protected by copyright. All rights reserved.Journal of Pineal Research 07/2014; · 7.81 Impact Factor
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ABSTRACT: Colon carcinogenesis is long known to be associated with ulcerative colitis (UC), a chronic gastrointestinal disorder. Various pre-clinical and clinical studies have shown that melatonin (MEL) has beneficial effects in cancer. However, elucidation of the detailed molecular mechanisms involved in MEL-mediated protection against the colon carcinogenesis deserves further investigation. The present study was aimed at deciphering the effect of MEL on autophagy and Nrf2 signaling pathways in a mouse model of colitis-associated colon carcinogenesis (CACC). For the induction of CACC, male Swiss Albino mice were administered a single ip injection of 20 mg 1, 2-dimethylhydrazine dihydrochloride (DMH)/kg bw, followed by 3 cycles of 3% w/v dextran sulfate sodium (DSS) in drinking water treatment initiated 1 wk after DMH injection. One week after the initiation of DSS treatment, MEL was administered at the dose of 1 mg/kg, bw, po for 8 and 18 wk. Mice were sacrificed at 10 and 20 wk after DMH injection. MEL treatment decreased the progression of CACC by down regulating the process of autophagy as revealed by the expression pattern of various autophagy markers such as Beclin-1, LC3B-II/LC3B-I ratio and p62. These findings were accompanied with the increased expression of Nrf2 and the associated antioxidant enzymes, NAD(P)H:quinone oxidoreductase (NQO-1) and heme oxygenase-1 (HO-1) in the colon of mice with CACC. MEL intervention reduced autophagy by ameliorating inflammation and oxidative stress in the colon of mice with CACC. We conclude that MEL treatment attenuates the progression of CACC in mice by modulating autophagy and Nrf2 signaling pathways.Molecular Carcinogenesis 01/2015; Ahead of print]. · 4.27 Impact Factor
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ABSTRACT: An ultra-performance liquid chromatography-quadrupole time-of-flight-based metabolomic approach was developed to study influence of salidroside, an anti-fatigue ingredient from Rhoiola rosea, on urinary metabolic profiling of rats to a single dose of 180 mg/kg per day. Unsupervised principal component analysis (PCA) and supervised orthogonal pre-projection to latent structures discriminate analysis (OPLS-DA) on metabolite profiling revealed obvious differentiation between the salidroside treated groups and controls in both positive and negative ion modes. Eleven urinary metabolites contributing to the differentiation were identified as anti-fatigue biomarkers: N-acetylserotonin, 2-Methoxyestrone 3-glucuronide, Taurine, Melatonin, Sorbitol, Geranyl diphosphate, Z-nucleotide, Cortisone, Dihydrocortisol, Sebacic acid, Pregnenolone sulfate. The physiological significance of these biomarkers is discussed. The work showed that metabolomics is a powerful tool in studying the anti-fatigue effects of natural compound salidroside on multiple targets in vivo.Journal of Pharmaceutical and Biomedical Analysis 02/2015; 105. · 2.83 Impact Factor