Stable free radicals as ubiquitous components of red wines

A stable ESR signal, centred at g = 2.0037 ± 0.0002, characterised by a single resonance and assignable to a free radical, was found in all the bottled red wines, both commercial and experimental, that we have examined. The radical concentration was calculated to be in the range of 5–82 nM. After exposure of the wines to air for a few minutes a two fold increase of the ESR signal, followed by a slow decrease with time, was observed. The intensity of ESR signal in experimental red wines, was found to increase with the ageing of the wines and was strictly correlated to the total content of polyphenols. The formation of semiquinone radicals of polyphenols is suggested as one possible mechanism leading to the presence of stable free radicals in red wines.     

Stable free radicals in ozone-damaged wheat leaves

Chlorophyll fluorescence measurements were performed on attached leaves of wheat plants (Triticum aestivum L. cv. Nandu) that were exposed to ambient air and to air supplemented with 80 and 120nmol mol^-1 ozone. Decreases in the “current photochemical capacity” were observed that were dependent on both the ozone concentration and duration of exposure. Electron paramagnetic resonance (EPR) spectra on freeze-dried samples from the same batches of plants showed the presence of an unidentified stable free radical, whose spectra had similarities to that of the ubisemiquinone radical. The intensity of this radical signal increased with the duration of ozone exposure in leaves that received an additional 120nmol mol^-1 ozone. In contrast, with exposure to air with 80nmol mol^-1 added ozone, there was little if any change in free radical signal intensity over the 4 week period of the experiment. The increase in intensity of the EPR signal occurred later than the chlorophyll fluorescence changes, which suggests that it is associated with permanent leaf damage.     

Stable free radicals in ozone-damaged wheat leaves

Chlorophyll fluorescence measurements were performed on attached leaves of wheat plants (Triticum aestivum L. cv. Nandu) that were exposed to ambient air and to air supplemented with 80 and 120nmol mol^-1 ozone. Decreases in the “current photochemical capacity” were observed that were dependent on both the ozone concentration and duration of exposure. Electron paramagnetic resonance (EPR) spectra on freeze-dried samples from the same batches of plants showed the presence of an unidentified stable free radical, whose spectra had similarities to that of the ubisemiquinone radical. The intensity of this radical signal increased with the duration of ozone exposure in leaves that received an additional 120nmol mol^-1 ozone. In contrast, with exposure to air with 80nmol mol^-1 added ozone, there was little if any change in free radical signal intensity over the 4 week period of the experiment. The increase in intensity of the EPR signal occurred later than the chlorophyll fluorescence changes, which suggests that it is associated with permanent leaf damage.     

Oxygen free radicals as pathogenic molecules in viral diseases.

Oxygen free radicals such as superoxide anion (O2-) were generated markedly in influenza virus-infected mouse lung, and these molecular species were identified as the potent pathogenic agents. This finding has many important implications for understanding viral pathogenesis: namely, the direct viral cytotoxicity (referred cytopathic effect) is only a fraction of several types of events induced by virus infection. The toxicity and reactivity of oxygen radicals, which are presumably generated in excessive amounts by the overreaction of the host's immune response against the organs or tissues in which viruses are replicating, may explain the mechanism of tissue injuries observed not only in influenza virus infection in mice, but also in other types of viral diseases in which immunological interactions are usually involved.     

Non-LTR retrotransposons (LINEs) as ubiquitous components of plant genomes

During the course of work aimed at isolating a rice gene from Oryza australiensis by PCR, the oligonucleotide primers used were found to generate a fragment that showed sequence homology to the endonuclease (EN) region of the maize non-LTR retrotransposon (LINE) Cin4. We carried out further PCRs using oligonucleotide primers that hybridized to these sequences, and found that they amplified several fragments, each with homology to the EN regions, from Oryza sativa cv. Nipponbare as well as O. australiensis. We mapped the approximate locations of two rice LINE homologues by screening clones in a YAC library made from a rice (O. sativa) genome, and found that each homologue was present in a low copy number apparently at nonspecific regions on rice chromosomes. We then carried out PCR using degenerate oligonucleotide primers which hybridized to the rice LINE homologues and Cin4 to ascertain whether LINE homologues are present in a variety of members of the plant kingdom, including angiosperms, gymnosperms, bracken, horsetail and liverwort. Cloning and nucleotide sequencing revealed that 53 clones obtained from 27 out of 33 plant species contained LINE homologues. In addition to these homologues, we identified four homologues with EN regions in the Arabidopsis thaliana genome by a computer search of databases. The nucleotide sequences of almost all the LINE homologues were greatly diverged, but the derived amino acid sequences were well conserved, and all contained glutamic acid and tyrosine residues at almost the same relative positions as in the the active site regions of AP (apurinic/apyrimidinic)-endonucleases. The EN regions in the LINE homologues from closely related plant species show a closer phylogenetic relationship, indicating that sequence divergence during vertical transmission has been a major influence upon the evolution of plant LINEs. Received: 13 July 1998 / Accepted: 13 October 1998     

Fatty acid compositions of commercial red wines

A determination was made of fatty acid compositions in twelve commercial red wines made from grapes differing in kind and vintage. Twelve fatty acids were identified, palmitic, myristic, and lauric acids being found predominant. Total acyls (32 approximately 81 nmol/100 ml) differed considerably. Changes in fatty acid constituents in must from grape berries and wines according to the process of manufacture were also examined.     

中国野生毛葡萄酒的香味成分分析

毛葡萄酒的香味成分由醇、酯、醛、酮等构成,3-甲基-1-丁醇是最主要的香气成分,分别占总量的34.1％（83-4-67）、34.12％（泰-12）、41.78％（商-24）;其单宁、总酚含量高,颜色深浓。毛葡萄种内3个系酒的香味成分总量、种类和各类成分的含量均存在明显差异。毛葡萄酒具有青草、烟熏、肥皂、焦糖及酸涩等香味特征。     

Phosphorus-containing iminoxyl and nitroxyl free radicals as promising spin labels

The structure and magnetic parameters of a series of nitroxyl and iminoxyl radicals differing in the position of the P atom with respect to the radical center were established by EPR. Free radicals were obtained in chemical and electrochemical reactions and also through UV irradiation with the use of spin traps. The phosphononitroxyls and phosphonoiminoxyls have proved quite sensitive to the milieu and temperature, which makes them promising spin labels for biological objects.     

Synthesis and biological evaluation of fused oxepinocoumarins as free radicals scavengers

Some fused dihydrooxepino[f]-, [g]-, and [h]coumarins were obtained from the ring-closing metathesis of the corresponding o-allyl-allyloxycoumarins under the treatment with the first generation Grubbs' catalyst. These compounds were tested in vitro for their antioxidant activity, and they present significant scavenging activity. They were also showed to inhibit in vitro soybean lipoxygenase.     

Synthesis and biological evaluation of fused oxepinocoumarins as free radicals scavengers

Some fused dihydrooxepino[f]-, [g]-, and [h]coumarins were obtained from the ring-closing metathesis of the corresponding o-allyl-allyloxycoumarins under the treatment with the first generation Grubbs’ catalyst. These compounds were tested in vitro for their antioxidant activity, and they present significant scavenging activity. They were also showed to inhibit in vitro soybean lipoxygenase.     

Hypochlorous acid as a precursor of free radicals in living systems

Hypochlorous acid (HOCl) is produced in the human body by the family of mammalian heme peroxidases, mainly by myeloperoxidase, which is secreted by neutrophils and monocytes at sites of inflammation. This review discusses the reactions that occur between HOCl and the major classes of biologically important molecules (amino acids, proteins, nucleotides, nucleic acids, carbohydrates, lipids, and inorganic substances) to form free radicals. The generation of such free radical intermediates by HOCl and other reactive halogen species is accompanied by the development of halogenative stress, which causes a number of socially important diseases, such as cardiovascular, neurodegenerative, infectious, and other diseases usually associated with inflammatory response and characterized by the appearance of biomarkers of myeloperoxidase and halogenative stress. Investigations aimed at elucidating the mechanisms regulating the activity of enzyme systems that are responsible for the production of reactive halogen species are a crucial step in opening possibilities for control of the development of the body’s inflammatory response.     

The role of free radicals in asbestos-induced diseases.

Asbestos exposure causes pulmonary fibrosis and malignant neoplasms by mechanisms that remain uncertain. In this review, we explore the evidence supporting the hypothesis that free radicals and other reactive oxygen species (ROS) are an important mechanism by which asbestos mediates tissue damage. There appears to be at least two principal mechanisms by which asbestos can induce ROS production; one operates in cell-free systems and the other involves mediation by phagocytic cells. Asbestos and other synthetic mineral fibers can generate free radicals in cell-free systems containing atmospheric oxygen. In particular, the hydroxyl radical often appears to be involved, and the iron content of the fibers has an important role in the generation of this reactive radical. However, asbestos also appears to catalyze electron transfer reactions that do not require iron. Iron chelators either inhibit or augment asbestos-catalyzed generation of the hydroxyl radical and/or pathological changes, depending on the chelator and the nature of the asbestos sample used. The second principal mechanism for asbestos-induced ROS generation involves the activation of phagocytic cells. A variety of mineral fibers have been shown to augment the release of reactive oxygen intermediates from phagocytic cells such as neutrophils and alveolar macrophages. The molecular mechanisms involved are unclear but may involve incomplete phagocytosis with subsequent oxidant release, stimulation of the phospholipase C pathway, and/or IgG-fragment receptor activation. Reactive oxygen species are important mediators of asbestos-induced toxicity to a number of pulmonary cells including alveolar macrophages, epithelial cells, mesothelial cells, and endothelial cells. Reactive oxygen species may contribute to the well-known synergistic effects of asbestos and cigarette smoke on the lung, and the reasons for this synergy are discussed. We conclude that there is strong evidence supporting the premise that reactive oxygen species and/or free radicals contribute to asbestos-induced and cigarette smoke/asbestos-induced lung injury and that strategies aimed at reducing the oxidant stress on pulmonary cells may attenuate the deleterious effects of asbestos.     

Biochemical aspects of free radicals

Toxic free radicals can be produced by many reactions required for the maintenance of normal metabolism and the production of energy in the cell. The reactivity of both primary and secondary radicals with biomolecules and in whole tissue systems is of interest, not only because of their importance in radiobiology but also because of the role these species play in toxicity and various disorders. Oxidant stress is known to increase the production of free radicals. In the presence of metals, especially iron, these radicals are converted into more damaging species. Trace elements play an important role in many systems that have evolved to deal with free radicals. The dietary status of the cell can affect the preventative antioxidant constituents of the cell. The chain-breaking antioxidant status can clearly be influenced by the dietary content of substances such as vitamins E and C.     

Modulation of fibroblast proliferation by oxygen free radicals.

The major unexplained phenomenon in fibrotic conditions is an increase in replicating fibroblasts. In this report we present evidence that oxygen free radicals can both stimulate and inhibit proliferation of cultured human fibroblasts, and that fibroblasts themselves release superoxide (O2.-) free radicals. Fibroblasts released O2.- in concentrations which stimulated proliferation, a finding confirmed by a dose-dependent inhibition of proliferation by free radical scavengers. Oxygen free radicals released by a host of agents may thus provide a very fast, specific and sensitive trigger for fibroblast proliferation. Prolonged stimulation may result in fibrosis, and agents which inhibit free radical release may have a role in the prevention of fibrosis.     

Eumelanins as free radicals trap and superoxide dismutase activities in Amphibia

• 1.1. Electron paramagnetic resonance (EPR) studies concerning the interaction between purified melanins and superoxide anion are reported; in addition the superoxide dismutase activities in the liver and brain of frog and rat were measured. Liver melanins considerably inhibit the EPR signal ascribed to superoxide anion.
• 2.2. The rat and frog brain show similar values of Cu, Zn SOD and Mn SOD. On the contrary, in frog liver, the specific activity of Cu, Zn SOD was significantly lower when compared to rat; no Mn SOD activity has been found in the frog liver.

     

UPLC analysis of free amino acids in wines: profiling of on-lees aged wines

The evolution of free amino acid (FAA) profiles intrinsic to on-lees aged white wines was determined by ultra performance liquid chromatography (UPLC?). On basis of the AccQ.Tag? method as a commercialized amino acid analysis solution for HPLC, a new protocol for dedicated amino acid analysis using 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) for pre-column derivatization was established by method transfer onto UPLC? conditions. Since AQC derivatives enable both fluorescence (AccQ.Tag? method) and UV detection, the performed method transfer additionally included changing to a more versatile UV detection. Emphasizing enhanced performance of UPLC?, the newly established protocol facilitated rapid and reliable separations of 24 amino acids within 23 min, hence proved to be superior compared to the original HPLC protocol due to significant improvements in resolution and reduced runtime. Applying UV detection enabled adequate quantifications of AQC amino acid derivatives at μM level (LOQs from 0.12 to 1.10 μM), thus proved sufficient sensitivity for amino acid profiling in wine samples. Moreover, this compiled methodology was successfully applied to monitor the changes of FAA concentrations in four distinct sets of on-lees aged white wines (fermented with different yeasts) at three progressing ripening periods, each (control, 3 and 6 months aging). For the control wines, the applied winery yeast significantly affected total FAA amounts (1450–1740 mg L^?1). During maturation, the proceeding yeast autolysis implied a rather complex impact on FAAs, yielding total FAA excretions up to 360 mg L^?1. However, the magnitude for increases of specific FAAs (up to +200%) highly depended on the individual amino acids as well as on the applied fermenting yeast. Given the overall complexity of yeast autolysis in winemaking, the application of efficient LC techniques such as UPLC? may indeed contribute as a valuable tool in wine research for product monitoring and characterization of intrinsic developments during wine maturation.     

Ion transport systems as targets of free radicals during ischemia reperfusion injury

Oxidative stress is a recognized pathogenic factor in ischemia/reperfusion injury (IRI). Iron induced generation of reactive oxygen species (ROS) in vitro reduces both the Na+K+-ATPase activity and Na+-Ca2+ exchanger of synaptosomal membranes, concomitantly with alteration of physical state of membranes. Oxidative insult also leads to the loss of ability of endoplasmic reticular membranes (ER) to sequester Ca2+ as well as to the increase of Ca2+ permeability. Furthermore, ROS induces both lipid peroxidation and lipid-independent modifications of membrane proteins. Acute in vivo ischemia alters kinetic parameters of Na+K+-ATPase affecting mainly the dephosphorylation step of ATPase cycle with parallel changes of Na+-Ca2+ exchanger and alterations of physical membrane environment. Subsequent reperfusion after ischemia is associated with decrease of immuno signal for PMCA 1 isoform in hippocampus. In addition, incubation of non-ischemic membranes with cytosol from ischemic hippocampus decreases level of PMCA 1 in non-ischemic tissues. Loss of PMCA 1 protein is partially protected both by calpain- and by non-specific protease inhibitors which suggest possible activation of proteases in the reperfusion period. On the other hand, ischemia does not affect the level of Ca2+ pump (SERCA 2b) and calreticulin of intracellular Ca2+ stores. However, IRI resulted in a decrease of IP3 receptor I and altered active Ca2+ accumulation into the ER. A non-specific alteration of physical properties of total membranes such as the oxidative modifications of proteins as well as the content of lipoperoxidation products can also be detected after IRI. ROS can alter physical and functional properties of neuronal membranes. We discuss our results suggesting that ischemia-induced disturbation of ion transport systems may participate in or follow delayed death of neurons after ischemia.     

Simple sequences are ubiquitous repetitive components of eukaryotic genomes

Simple sequences are stretches of DNA which consist of only one, or a few tandemly repeated nucleotides, for example poly (dA) X poly (dT) or poly (dG-dT) X poly (dC-dA). These two types of simple sequence have been shown to be repetitive and interspersed in many eukaryotic genomes. Several other types have been found by sequencing eukaryotic DNA. In this report we have undertaken a systematical survey for simple sequences. We hybridized synthetical simple sequence DNA to genome blots of phylogenetically different organisms. We found that many, probably even all possible types of simple sequence are repetitive components of eukaryotic genomes. We propose therefore that they arise by common mechanisms namely slippage replication and unequal crossover and that they might have no general function with regards to gene expression. This latter inference is supported by the fact that we have detected simple sequences only in the metabolically inactive micronucleus of the protozoan Stylonychia, but not in the metabolically active macronucleus which is derived from the micronucleus by chromosome diminution.     

Synthesis of purine homo-N-nucleosides modified with coumarins as free radicals scavengers*

Cross metathesis (CM) of 9-butenylpurines with 4-butenyloxycoumarin in the presence of Grubbs 2nd generation catalyst under MW irradiation resulted to conjugated compounds containing homo-N-nucleosides and coumarins. Analogous derivatives received by the CM reaction of 9-butenyl-6-piperidinylpurine with 6- or 7-butenyloxycoumarins, allyloxycoumarins or coumarinyl acrylate. These compounds were tested in vitro for their antioxidant activity and they present significant scavenging activity. The presence of a pentenyloxy moiety, the attachment position on coumarin ring as well as a purine homo-N-nucleoside group are considered as important structural features.