Oxidation of dihydronicotinamide adenine dinucleotide by a flavin derivative of polyethylenimine

A modified Polyethylenimine has been prepared that has riboflavin attached to it, as well as hydrophobic groups. The catalytic efficiency toward oxidation of6 dihydronicotinamide adenine dinucleotide (NADH) by this flavopolymer is more than 100-fold greater than observed with small-molecular riboflavin. The products of the reaction in aerobic solution have been established to be the oxidized nicotinamide NAD⁺ and hydrogen peroxide. The kinetics fit a Michaelis-Menten mechanism. Thus, the versatility of modified polyethylenimines as catalysts has been extended from hydrolyses and decarboxylations to oxidation-reduction reactions also.     

Flash photolysis of flavins. V. Oxidation and Disproportionation of flavin radicals

Flash photolysis techniques have been utilized to investigate the reactivity patterns of flavin radical species. Rate constants for disproportionation were found to be la the following order: lumiflavin>FMN>FAD and neutral radicals>anionic radicals. Neutral flavin radicals react with oxygen at a rate which is at least 10⁴ times slower than the anionic species. No evidence for an intermediate complex or adduct is obtained in this reaction. The pK values for the ionization of the neutral flavin radicals are in the order FAD>FMN>riboflavin=limiflavin. The rates of reaction of ferricyanide with flavin radicals are essentially independent of pH, whereas benzoquinone reacts slightly more slowly (5 times) with the neutral flavin radical than with the anionic form. Cytochromec reacts at least ten times more slowly with flavin radicals than does ferricyanide.     

Indices of oxidative stress. 2. Lipid peroxides

Two methods of the determination of lipid peroxidation products have been compared which are based on Fe(II) oxidation by them at acid pH values in the presence of xylenol orange which binds Fe(III) have been compared. The first method uses cumene hydropeoxide as an internal standard. In the second one, lipid peroxides are previously reduced by triphenylphosphine and these substances content is measured as a difference of the production of complexes with xylenol orange and iron ions in the control (with reduction) and experimental sample (without reduction). The optimization of measurement conditions is described. The levels of lipid peroxides in goldfish tissues assayed simultaneously by two methods were similar. The method with cumene hydroperoxide needs less amounts of biological material; moreover, there is no necessity in a calibration curve. Effects of hyperoxia on lipid peroxide levels in goldfish tissues were studied with the cumene method. Within the first hours of hyperoxia this index increased 13-times in the liver and 2-times in the brain and muscle. The further exposure rebounded this parameter to the initial level. Levels of lipid peroxides positively correlated with levels of end products of lipid peroxidation (thiobarbiturate acid reactive substances) in the goldfish tissues. The method of quantification of lipid peroxides with cumene is recommended for wide using in biological investigations.     

Cyanide Formation from Oxidation of Glycine by a Pseudomonas Species

With whole cells of a hydrogen cyanide-producing bacterium strain C, of the genus Pseudomonas, it was found that the oxygen necessary for the oxidation of glycine to cyanide could be replaced by various artificial electron acceptors. The order of reactivity was: oxygen > phenazine methosulphate > methylene blue > 2,6-dichlorophenolindophenol > ferricyanide. Cyanide production was inhibited by pyrrolnitrin, a well-known inhibitor of many flavine enzymes. The molar ratio of added glycine to cyanide produced was found to be 1.09. With whole bacteria the apparent K(m) (glycine) for the cyanide production was found to be 5.0 x 10(-4) M.     

Prostaglandin biosynthesis can be triggered by lipid peroxides.

Studies of ferriheme cyclooxygenase, using two different assay systems, show that a variety of peroxides can trigger a rapid acceleration of cyclooxygenase activity to produce prostaglandins. Lipid hydroperoxides formed by lipoxygenase were the most potent activators tested, followed by prostaglandin G₂, which was slightly less potent. Peroxides nonspeciflcally generated during arachidonate autoxidation were as potent as the enzymatically formed lipid peroxides. These findings have important implications for cell function since any process which generates peroxides may activate the cyclooxygenase. Thus the balance between formation and removal of cellular lipid peroxides sets a peroxide tone that can regulate the rate of prostaglandin formation in cells.     

Clustering of apoptotic cells via bystander killing by peroxides.

Clustering of apoptotic cells is a characteristic of many developing or renewing systems, suggesting that apoptotic cells kill bystanders. Bystander killing can be triggered experimentally by inducing apoptosis in single cells and may be based on the exchange of as yet unidentified chemical cell death signals between nearby cells without the need for cell-to-cell communication via gap junctions. Here we demonstrate that apoptotic cell clusters occurred spontaneously, after serum deprivation or p53 transfection in cell monolayers in vitro. Clustering was apparently induced through bystander killing by primary apoptotic cells. Catalase, a peroxide scavenger, suppressed bystander killing, suggesting that hydrogen peroxide generated by apoptotic cells is the death signal. Although p53 expression increased the number of apoptoses, clustering was found to be similar around apoptotic cells whether or not p53 was expressed, indicating that there is no specific p53 contribution to bystander killing. Bystander killing through peroxides emitted by apoptotic cells may propagate tissue injury in different pathological situations and be relevant in chemo-, gamma-ray, and gene therapy of cancer.     

Selective Inhibition of Ammonium Oxidation and Nitrification-Linked N2O Formation by Methyl Fluoride and Dimethyl Ether

Methyl fluoride (CH₃F) and dimethyl ether (DME) inhibited nitrification in washed-cell suspensions of Nitrosomonas europaea and in a variety of oxygenated soils and sediments. Headspace additions of CH₃F (10% [vol/vol]) and DME (25% [vol/vol]) fully inhibited NO₂^- and N₂O production from NH₄⁺ in incubations of N. europaea, while lower concentrations of these gases resulted in partial inhibition. Oxidation of hydroxylamine (NH₂OH) by N. europaea and oxidation of NO₂^- by a Nitrobacter sp. were unaffected by CH₃F or DME. In nitrifying soils, CH₃F and DME inhibited N₂O production. In field experiments with surface flux chambers and intact cores, CH₃F reduced the release of N₂O from soils to the atmosphere by 20- to 30-fold. Inhibition by CH₃F also resulted in decreased NO₃^- + NO₂^- levels and increased NH₄⁺ levels in soils. CH₃F did not affect patterns of dissimilatory nitrate reduction to ammonia in cell suspensions of a nitrate-respiring bacterium, nor did it affect N₂O metabolism in denitrifying soils. CH₃F and DME will be useful in discriminating N₂O production via nitrification and denitrification when both processes occur and in decoupling these processes by blocking NO₂^- and NO₃^- production.     

Formation of HCN and its chlorination to ClCN by stimulated human neutrophils--2. Oxidation of thiocyanate as a source of HCN

Leucocytes challenged by Staphylococcus epidermidis or stimulated by phorbol myristate acetate (PMA) produce cyanide from thiocyanate. The amount of H14CN formed depends on KS14CN concentration and is enhanced by pretreatment of phagocytosed bacteria with penicillin or by adding amine-taurine to the medium of PMA-stimulated neutrophils. The reaction of taurine chloramine or chlorinated Staphylococcus epidermidis (containing N-Cl groups) with thiocyanate results in HCN formation. At higher concentration of chloramine cyanogen chloride is formed. Cyanide is chlorinated by PMA-stimulated neutrophils and this process is significantly enhanced by exogenous taurine and inhibited by 3-amino 1,2,4-triazole. It is conceivable that oxidation of thiocyanate to HCN and chlorination of HCN to ClCN is mediated by the chlorinating species (taurine chloramine) produced by stimulated neutrophils.