Oxidative stress occurring in cells is a consequence of an excessive activity of reactive oxygen forms, resulting from an imbalance between the release of free oxygen radicals and their removal from the cell by antioxidant systems. 90% of reactive oxygen radicals emerge in mitochondrial respiratory chain during an incomplete four-electron oxygen reduction. The remaining 10% originate from different reactions occurring in the cell. The established compounds are characterised by a short half-life and are highly reactive. Sparse quantities of free oxygen radicals have a positive effect on cell functions. Oxidative stress leads to damage in cellular membranes, enzymatic and non-enzymatic proteins, as well as DNA. Therapy with antioxidants as exogenous dietary supplements aims at preventing or reducing the risk of development of diseases involving the presence of the oxygen radicals. Whether the antioxidant therapy will bring positive or negative effects depends on numerous factors that need to be considered before their inclusion in the applied treatment.
Jelena Pantic Bisevac, Mirjana Djukic, Ivan Stanojevic, Ivana Stevanovic, Zeljko Mijuskovic, Ana Djuric, Borko Gobeljic, Tatjana Banovic and Danilo Vojvodic
Background: Overproduction of free radicals accompanied with their insufficient removal/neutralization by antioxidative defense system impairs redox hemostasis in living organisms. Oxidative stress has been shown to be involved in all the stages of carcinogenesis and malignant melanocyte transformation. The aim of this study was to examine association between oxidative stress development and different stages of melanoma. Methods: The measured oxidative stress parameters included: superoxide anion radical, total and manganese superoxide dismutase, catalase and malondialdehyde. Oxidative stress parameters were measured spectrophotometrically in serum samples from melanoma patients (n=72) and healthy control subjects (n=30). Patients were classified according to AJCC clinical stage. Results: Average superoxide anion and malondialdehyde concentrations were significantly higher in melanoma patients than in control group, with the highest value of superoxide anion in stage III, while malondialdehyde highest value was in stage IV. The activity of total and manganese superoxide dismutase was insignificantly higher in melanoma patients than in control group, while catalase activity was significantly higher. The highest activity of total activity of manganese superoxide dismutase was in stage IV. Catalase activity was increasing with the disease progression achieving the maximum in stage III. Conclusion: Results of our study suggest that melanoma is oxidative stress associated disease, as well as deteriorated cell functioning at mitochondrial level.
Coffee is a rich source of dietary antioxidants which protects the human body against the effects of dangerous free radicals. The aim of this study was to determine and compare the antioxidant activity, content of total phenols and flavonoids in selected types of coffee with respect to the way of their processing. The individual coffees were investigated with regard to their origin and composition. The antioxidant effects were determined by the 2,2-diphenyl-1-picryl-hydrazyl (DPPH) radical scavenging assay. The content of total phenols was analysed by the Folin-Ciocalteu method and the content of flavonoids in the coffee extracts was determined by a colorimetric method. The highest antioxidant activity was exhibited by the extract of unroasted ground 100 % green coffee Arabica (89.55 %), and the high scavenging of free radicals was achieved also by the extracts of roasted ground 100 % coffees Arabica and Robusta. The highest levels of total phenols (77.54 µg.ml−1) and flavonoids (1.74 µg.ml−1) were measured in the extract of unroasted ground 100 % green coffee Arabica. High levels of these substances were found also in extracts of roasted ground 100 % coffees Arabica and Robusta. The lowest levels of total phenols (31.24 µg.ml−1) and flavonoids (0.91 µg.ml−1) were detected in the extract of instant coffee Arabica. The processing of coffee by roasting decreased the level of the investigated antioxidant components but considerably improved the taste and aroma, the properties that make coffee one of the most popular drinks in the world.
Amir Shahbazfar, Payman Zare, Hemn Mohammadpour and Hossein Tayefi-Nasrabadi
Effects of different concentrations of artemisinin and artemisinin-iron combination treatment on Madin Darby Canine Kidney (MDCK) cells
Artemisinin is a sesquitrepenelactone with an endoperoxide bridge. It is a naturally occurring substance from Artemisia species plants. Artemisia species have been used in oriental medicine for centuries to treat malaria, gastrointestinal helminthosia, diarrhea, and as an antipyretic and sedative agent. Antileishmanial activity of the plants has been announced a few years ago. Dogs are the most important reservoir of leishmaniasis in some parts of the world. To use it as an antileishmanial drug in dogs, its side effects on different organs, among them the kidney as the organ of elimination have to be elucidated. Artemisinin with different concentrations (0.15, 0.3, 0.6 and 1.2 μg/ml) was added to the culture of MDCK (Madin darby canine kidney) cells with and without iron (86 μg/dl). All the changes were controlled and photographed every 12 hours using an invert microscope. After 60 hours, supernatants and cell extracts were examined for LDH (lactate dehydrogenase) concentration and total protein. Also TBARS (thiobarbituric acid reactive substances) test was performed on cell extracts. Some microscopic slides were prepared from the cells and stained with hematoxylin-eosin for microscopic exams. Biochemical parameters showed cellular reaction and injury in a concentration dependent manner. Cell injury was more severe in the iron-added groups. Microscopic exams showed cell and nuclear swelling, granular degeneration, vacuole and vesicle formation, cellular detachment, piknosis, karyorrhexis, cellular necrosis and inhibition of new mitosis. On using the drug for leishmaniasis treatment in the dog, it should be done with caution and supervision.
The popular pharmaceutical base used in pharmacy – vaselinum flavum – was studied by an X-band (9.3 GHz) EPR spectrometer in the range of microwave power of 2.2–70 mW. The samples were sterilized in hot air oven at temperatures: 160°C (120 min), 170°C (60 min), and 180°C (30 min). The aim of this work was to determine properties and free radical concentrations in vaselinum flavum thermally sterilized at different conditions. The changes in free radical system in vaselinum flavum during storage were analyzed. Free radicals were found in all the heated samples. The lowest free radical concentration was obtained for vaselinum flavum heated at 180°C for 30 min; so these parameters are proposed for the thermal sterilization of this pharmaceutical base. Interactions with oxygen decreased free radical concentration in vaselinum flavum during storage. Strong quenching of free radicals in vaselinum flavum was observed after 2 days for the samples sterilized at temperatures 160 and 180°C. Such an effect for vaselinum flavum heated at temperature 170°C was observed later, 13 days after sterilization. Fast spin-lattice relaxation processes exist in thermally sterilized vaselinum flavum. The EPR lines of heated vaselinum flavum were homogeneously broadened. EPR spectroscopy and its use for examining the thermal sterilization process in pharmacy was confirmed.
Free radicals in synthetic melanin and melanin from Sepia officinalis were studied by electron paramagnetic resonance (EPR) spectroscopy. The effect of time of ultraviolet (UV) irradiation on free radicals in these melanins was tested. The samples were exposed to UV during 15, 30, and 60 minutes. EPR spectra were measured with microwaves from an X-band (9.3 GHz) in the range of microwave power of 2.2–70 mW. The performed EPR examinations indicate that high concentrations (~1021–1022 spin/g) of o-semiquinone free radicals with g factors of 2.0039–2.0045 exist in all the tested samples. For nonirradiated samples, free radical concentration was higher in natural melanin than in synthetic melanin. UV irradiation caused the increase of free radical concentrations in synthetic melanin samples and this effect depends on the time of irradiation. The largest free radical formation in the both melanins was obtained for 60 min of UV irradiation. Free radical concentrations after the UV irradiation of melanins during 30 min were lower than during irradiation by 15 min, and probably this effect was the result of recombination of the radiatively formed free radicals. EPR lines of the tested samples broadened with increasing microwave power, so these lines were homogeneously broadened. The two types of melanins differed in the time of spin-lattice relaxation processes. Slower spin-lattice relaxation processes exist in melanin from Sepia officinalis than in synthetic melanin. UV irradiation did not change the time of spin-lattice relaxation processes in the tested melanins. The performed studies confirmed the usefulness of EPR spectroscopy in cosmetology and medicine.
Radmila Radojevic-Popovic, Tamara Nikolic, Isidora Stojic, Jovana Jeremic, Ivan Srejovic, Goran Pesic and Vladimir Jakovljevic
The effect of scuba diving on ROS production and oxidative stress compared to that of other recreational activities is still poorly understood. The aim of this study was to assess the influence of different types of physical activity on the redox status of scuba divers by testing the pro- and anti-oxidative parameters immediately before and after different types of physical load. The prevalence study included 10 professional police divers. All examinees were male, 32 ± 5.1 years of age, well-trained, and with a minimum of five to a maximum of 20 years of diving experience. The study was divided into three experimental protocols: 1) an exercise test (at atmospheric pressure), 2) an at sea dive (30 meters for 30 minutes), and 3) a dive into river current (10 meters for 30 minutes). Immediately before and after the load test of the divers at atmospheric pressure and immediately before and after the dive, blood samples were taken to determine the values of the following pro-oxidant markers: O2−, H2O2, NO2− and TBARS, as well as antioxidant enzymes (SOD and CAT). A comparison of the results before and after physical activity for all three protocols revealed a significant increase in values for NO2−, O2−, H2O2 and CAT after physical activity. It can be concluded that the values of all oxidative stress markers depend on the season of the year in which the research is conducted or on the frequency of dives and degree of physical exertion during this period of the year.
Russel Reiter, Sergio Paredes, Ahmet Korkmaz, Mei-Jie Jou and Dun-Xian Tan
Melatonin combats molecular terrorism at the mitochondrial level
The intracellular environmental is a hostile one. Free radicals and related oxygen and nitrogen-based oxidizing agents persistently pulverize and damage molecules in the vicinity of where they are formed. The mitochondria especially are subjected to frequent and abundant oxidative abuse. The carnage that is left in the wake of these oxygen and nitrogen-related reactants is referred to as oxidative damage or oxidative stress. When mitochondrial electron transport complex inhibitors are used, e.g., rotenone, 1-methyl-1-phenyl-1,2,3,6-tetrahydropyridine, 3-nitropropionic acid or cyanide, pandemonium breaks loose within mitochondria as electron leakage leads to the generation of massive amounts of free radicals and related toxicants. The resulting oxidative stress initiates a series of events that leads to cellular apoptosis. To alleviate mitochondrial destruction and the associated cellular implosion, the cell has at its disposal a variety of free radical scavengers and antioxidants. Among these are melatonin and its metabolites. While melatonin stimulates several antioxidative enzymes it, as well as its metabolites (cyclic 3-hydroxymelatonin, N1-acetyl-N2-formyl-5-methoxykynuramine and N1-acetyl-5-methoxykynuramine), likewise effectively neutralize free radicals. The resulting cascade of reactions greatly magnifies melatonin's efficacy in reducing oxidative stress and apoptosis even in the presence of mitochondrial electron transport inhibitors. The actions of melatonin at the mitochondrial level are a consequence of melatonin and/or any of its metabolites. Thus, the molecular terrorism meted out by reactive oxygen and nitrogen species is held in check by melatonin and its derivatives.
Ryszard Krzyminiewski, Zdzislaw Kruczynski, Aleksander Stepien and Bernadeta Dobosz
Spin traps in the detection of free radicals in the blood of patients with ischemia
Electron Paramagnetic Resonance and a nitrosobenzene spin trap were used to investigate free radicals in the human blood after angioplasty treatment. The nitrosobenzene anion radical was determined using EPR measurements and quantum-mechanical calculations. Differences were observed in the concentration of free radicals before and after angioplasty treatment. These results were compared with myocardium damage parameters (CPK, MB and TnT).
Henrietta Ogbunugafor, Miriam Igwo-Ezikpe, Innocent Igwilo, Nwaorah Ozumba, Sunday Adenekan, Chidozie Ugochukwu, Obumuneme Onyekwelu and Anthony Ekechi
Background: The pathogenesis of reactive oxygen species (ROS) linked chronic diseases such as hypertension, rheumatoid arthritis, and diabetes has warranted the intensive search for plants with antioxidant properties.
Aim: The in vitro and in vivo antioxidant properties of ethanolic leaves extract of Moringa oleifera Lam (Moringaceae) and its effect on lipid indices in male albino rats were investigated.
Material and Methods: The in vitro antioxidant properties of the extract were determined; while the invivo studies involved three groups of five male rats orally administered with extract [1 mL (200 mg/kg- 1)], alpha-tocopherol [1 mL (500 mg/kg-1)] and 1 mL 5 % Tween 20 (control) respectively for 21 days. The effects on enzymatic antioxidants and lipid indices were evaluated.
Results: Hydrogen peroxide scavenging ability of the extract was significantly (p<0.05) higher than that of alpha-tocopherol. Superoxide dismutase and catalase activities were increased, while triacylglycerides and very low density lipoprotein were significantly (p<0.05) lowered by the extract. Malonaldehyde levels were not significantly (p<0.05) different in the extract, alpha-tocopherol and control treated groups.
Conclusion: The extract had appreciable phenol content and exhibited in vitro antioxidant properties. The extract also had modulating effects on enzymatic antioxidants, showed lipid-lowering ability and caused changes in types of lipoproteins found in serum.