Nitric Oxide Reversibly Suppresses Xanthine Oxidase Activity

The effects of nitric oxide (NO) on xanthine oxidase (XOD) activity and the site(s) of the redox center(s) affected were investigated. XOD activity was determined by superoxide (O₂^-) generation and uric acid formation. NO reversibly and dose-dependently suppressed XOD activity in both determination methods. The suppression interval also disclosed a dose-dependent prolongation. The suppression occurred irrespective of the presence or absence of xanthine; indicating that the reaction product of NO and O₂^-, peroxynitrite, is not responsible for the suppression. Application of synthesized peroxynitrite did not affect XOD activity up to 2 μM. Methylene blue, which is an electron acceptor from Fe/S center, prevented the NO-induced inactivation. The results indicate that NO suppresses XOD activity through reversible alteration of the flavin prosthetic site.     

The correlation between active oxygens scavenging and antioxidative effects of flavonoids

The abilities of 15 flavonoids as a scavenger of active oxygens (hydroxyl radical and superoxide anion) were studied. Hydroxyl radical (^.OH) was generated by the Fenton system, and assayed by the determination of methanesulfinic acid (MSA) formed from the reaction of dimethyl sulfoxide (DMSO) with ^.OH. (+)-Catechin, (−)-epicatechin, 7,8-dihydroxy flavone, and rutin showed the ^.OH scavenging effect 100–300 times superior to that of mannitol, a typical ^.OH scavenger. The other flavonoids showed no ^.OH scavenging effect at their concentrations up to 50 μM. Baicalein, quercetin, morin, and myricetin unexpectedly increased the ^.OH production in the Fenton system. The flavonoids tested now, except monohydroxy flavones, were more or less inhibitive to the superoxide anion (O₂) generation in the hypoxanthine-xanthine oxidase system. A great part of this inhibitory effect was likely owing to suppression of xanthine oxidase activity by the flavonoids. The flavonoids, which scavenged ^.OH or O₂⁻, were necessarily antioxidants to the peroxidation of methyl linoleate. However, there was a type of flavonoid such as morin, which have neither ^.OH nor O₂⁻ scavenging effect, but was a strong antioxidant.     

Bipyridylium quaternary salts and related compounds. V. Pulse radiolysis studies of the reaction of paraquat radical with oxygen. Implications for the mode of action of bipyridyl herbicides

1. Rate constants for reduction of paraquat ion (1,1′-dimethyl-4,4′-bipyridy-lium, PQ²⁺) to paraquat radical (PQ⁺·) by e⁻_aq and CO₂⁻· have been measured by pulse radiolysis. Reduction by e⁻_aq is diffusion controlled (k = 8.4·10¹⁰ M⁻¹·s⁻¹) and reduction by CO₂⁻· is also very fast k = 1.5·10¹⁰ M⁻¹·s⁻¹).

2. The reaction of paraquat radical with oxygen has been analysed to give rate constants of 7.7·10⁸ M⁻¹·s⁻¹ and 6.5·10⁸ M⁻¹·s⁻¹ for the reactions of paraquat radical with O₂ and O₂⁻·, respectively. The similarity in these rate constants is in marked contrast to the difference in redox potentials of O₂ and O₂⁻· (− 0.59 V and + 1.12 V, respectively).

3. These rate constants, together with that for the self-reaction of O₂⁻·, have been used to calculate the steady-state concentration of O₂⁻· under conditions thought to apply at the site of reduction of paraquat in the plant cell. On the basis of these calculations the decay of O₂⁻· appears to be governed almost entirely by its self-reaction, and the concentration 5 μm away from the thylakoid is still 90% of that at the thylakoid itself. Thus, O₂⁻· persists long enough to diffuse as far as the chloroplast envelope and tonoplast, which are the first structures to be damaged by paraquat treatment. O₂⁻· is therefore sufficiently long-lived to be a candidate for the phytotoxic product formed by paraquat in plants.     

Crystal and microparticle effects on MDCK cell superoxide production: oxalate-specific mitochondrial membrane potential changes

We have previously shown that crystals of calcium oxalate (COM) elicit a superoxide (O₂⁻) response from mitochondria. We have now investigated: (i) if other microparticles can elicit the same response, (ii) if processing of crystals is involved, and (iii) at what level of mitochondrial function oxalate acts. O₂⁻ was measured in digitonin-permeabilized MDCK cells by lucigenin (10 μM) chemiluminescence. [¹⁴C]-COM dissociation was examined with or without EDTA and employing alternative chelators. Whereas mitochondrial O₂⁻ in COM-treated cells was three- to fourfold enhanced compared to controls, other particulates (uric acid, zymosan, and latex beads) either did not increase O₂⁻ or were much less effective (hydroxyapatite +50%, p < 0.01), with all at 28 μg/cm². Free oxalate (750 μM), at the level released from COM with EDTA (1 mM), increased O₂⁻ (+50%, p < 0.01). Omitting EDTA abrogated this signal, which was restored completely by EGTA and partially by ascorbate, but not by desferrioxamine or citrate. Omission of phosphate abrogated O₂⁻, implicating phosphate-dependent mitochondrial dicarboxylate transport. COM caused a time-related increase in the mitochondrial membrane potential (Δψ_m) measured using TMRM fluorescence and confocal microscopy. Application of COM to Fura 2-loaded cells induced rapid, large-amplitude cytosolic Ca²⁺ transients, which were inhibited by thapsigargin, indicating that COM induces release of Ca²⁺ from internal stores. Thus, COM-induced mitochondrial O₂⁻ requires the release of free oxalate and contributes to a synergistic response. Intracellular dissociation of COM and the mitochondrial dicarboxylate transporter are important in O₂⁻ production, which is probably regulated by Δψ_m.     

Trehalose: Protector of antioxidant enzymes or reactive oxygen species scavenger under heat stress?

Trehalose is known to protect membranes and macromolecules. Its accumulation has been implicated in allowing plants to tolerate stress, including heat-shock. However, under heat-shock, it is not clear whether trehalose eliminates reactive oxygen species (ROS) directly or indirectly by protecting antioxidant enzymes. In this study, we initially examined the effects of trehalose on the activities of key antioxidant enzymes, including superoxide dismutases (SODs), ascorbate catalases (CATs), and ascorbate peroxidases (APX) from wheat (Triticum aestivum L.), and then measured the ability of trehalose to scavenge hydrogen peroxide (H₂O₂) and superoxide anions (O₂⁻). Our results indicated that trehalose protected SOD activity slightly. However, it inhibited CAT and APX activities under heat stress, with a little protection of CAT activity (only about 7% promotion) at 22 °C. Moreover, trehalose scavenged H₂O₂ and O₂⁻ greatly in a concentration-dependent manner, reaching the maximal scavenging H₂O₂ rate of 95% and O₂⁻ rate of 78%, respectively, at 50 mM trehalose. These results suggest that trehalose plays a direct role in eliminating H₂O₂ and O₂⁻ in wheat under heat stress.     

Examination of the Inhibitory Effect of Norathyriol in Formylmethionyl-Leucyl-Phenylalanine-Induced Respiratory Burst in Rat Neutrophils

Norathyriol, aglycone of a xanthone C-glycoside mangiferin isolated from Tripterospermum lanceolatum, concentration dependently inhibited the formylmethionyl-leucyl-phenylalanine (fMLP)-induced superoxide anion (O₂^˙−) generation and O₂ consumption in rat neutrophils. In cell-free oxygen radical generating system, norathyriol inhibited the O₂^˙− generation during dihydroxyfumaric acid (DHF) autoxidation and in hypoxanthine-xanthine oxidase system. fMLP-induced transient elevation of [Ca²⁺]_i and the formation of inositol trisphosphate (IP₃) were significantly inhibited by norathyriol (30 μM) (about 30 and 46% inhibition, respectively). Norathyriol concentration dependently suppressed the neutrophil cytosolic phospholipase C (PLC). In contrast with the marked attenuation of fMLP-induced protein tyrosine phosphorylation (about 70% inhibition at 10 μM norathyriol), norathyriol only slightly modulated the phospholipase D (PLD) activity as determined by the formation of phosphatidic acid (PA) and, in the presence of ethanol, phosphatidylethanol (PEt). Norathyriol did not modulate the intracellular cyclic AMP level. In the presence of NADPH, the phorbol 12-myristate 13-acetate (PMA)-activated particulate NADPH oxidase activity was suppressed by norathyriol in a concentration-dependent manner and the inhibition was noncompetitive with respect to NADPH. Norathyriol inhibited the iodonitrotetrazolium violet (INT) reduction in arachidonic acid (AA)-activated cell-free NADPH oxidase system at the same concentration range as those used in the suppression of PMA-activated particulate NADPH oxidase activity. Taken together, these results suggest that the scavenging ability of norathyriol contributes to the reduction of generated O₂^˙−, however, the inhibition of O₂^˙− generation from neutrophils by norathyriol is attributed to the blockade of PLC pathway, the attenuation of protein tyrosine phosphorylation, and to the suppression of NADPH oxidase through the interruption of electrons transport.     

A new type of O(2)(-)-generating tool for oxidative stress studies by remodeling neutrophil NADPH oxidase

The effects of reactive oxygen species on cells have attracted much attention in relation to redox regulation and oxidative stress-related diseases. Superoxide (O₂⁻) is the reactive oxygen species primarily formed in biological systems. However, no convenient O₂⁻-generating device has been available for use in cell or tissue culture. The neutrophil NADPH oxidase, a professional enzyme for killing bacteria, has a high ability to produce O₂⁻. However, the cell-free activation process requires several protein factors and an anionic amphiphile, and moreover, the activation is transient. To utilize the enzyme as an O₂⁻ generator, we improved the cell-free activation method by remodeling regulatory components, optimizing lipid composition, and modifying the mixing conditions. We established a new method to produce an active enzyme that is stable, efficient, and preservable. As an application, we examined the effect of the device on cultured HEK293 cells and observed that it caused cell death. This system has several advantages over the xanthine oxidase system often used. The new device will be useful for studies of oxidative stress and related diseases.     

Direct measurement of oscillatory generation of superoxide anions by single phagocytes

Phagocytic cells such as neutrophils generate superoxide anions (O₂⁻) within phagocytic vacuoles for killing and digesting microorganisms. Here we report the simultaneous observation of morphological changes and O₂⁻ generation in single phagocytic cells during phagocytosis. Point stimulation of a cell by contact with an opsonized microelectrode at the cell surface induced significant deformation to engulf the electrode, and also induced the O₂⁻ generation which was measured by the electrode. Periodic fluctuations in the magnitude of the O₂⁻ generation were observed in the time course. These oscillations may be caused by metabolic regulation of the formation of NADPH, which is the substrate for the O₂⁻ generation.     

Metal ion-coordinating properties of imidazole and derivatives in aqueous solution: interrelation between complex stability and ligand basicity

The stability constants of the 1:1 complexes formed between Mg²⁺, Ca²⁺, Sr²⁺, Ba²⁺, Mn²⁺, Co²⁺, Ni²⁺, Cu²⁺, Zn²⁺ or Cd²⁺ (M²⁺) and the simple, sterically unhindered imidazole-type ligands, imidazole, 1-methylimidazole, 5-chloro-1-methylimidazole, N-(2,3,5,6-tetrafluorophenyl)imidazole or 4′-(imidazol-1-yl)acetophenone (L), were determined by potentiometric pH titrations in aqueous solution (25°C; I = 0.5 M, NaNO₃). The construction of log K_ML^M versus pK_HL^H plots results in straight lines; the equations for the least-squares lines are calculated and listed. These data allow calculation of the expected stability constant for a complex of any imidazole-type ligand, provided its pK_HL^H value (in the pK_a range 4–8) is known. For the stabilities of Fe²⁺ complexes with imidazole-type ligands an estimation procedure is provided. It is shown further that the complex formation between 1-methylbenzimidazole (MBI) and Mn²⁺, Ni²⁺, Cu²⁺ or Zn²⁺ is s sterically hindered, i.e. the data points for these M(MBI)²⁺ complexes do not fall on the straight lines defined by the imidazole-type ligands.     

Mechanisms of H2O2-induced oxidative stress in endothelial cells

Hydrogen peroxide, produced by inflammatory and vascular cells, induces oxidative stress that may contribute to endothelial dysfunction. In smooth muscle cells, H₂O₂ induces production of O₂⁻ by activating NADPH oxidase. However, the mechanisms whereby H₂O₂ induces oxidative stress in endothelial cells are poorly understood. We examined the effects of H₂O₂ on O₂⁻ levels on porcine aortic endothelial cells (PAEC). Treatment with 60 μmol/L H₂O₂ markedly increased intracellular O₂⁻ levels (determined by conversion of dihydroethidium to hydroxyethidium) and produced cytotoxicity (determined by propidium iodide staining) in PAEC. Overexpression of human manganese superoxide dismutase in PAEC reduced O₂⁻ levels and attenuated cytotoxicity resulting from treatment with H₂O₂. L-NAME, an inhibitor of nitric oxide synthase (NOS), and apocynin, an inhibitor of NADPH oxidase, reduced O₂⁻ levels in PAEC treated with H₂O₂, suggesting that both NOS and NADPH oxidase contribute to H₂O₂-induced O₂⁻ in PAEC. Inhibition of NADPH oxidase using apocynin and NOS rescue with L-sepiapterin together reduced O₂⁻ levels in PAEC treated with H₂O₂ to control levels. This suggests interaction-distinct NOS and NADPH oxidase pathways to superoxide. We conclude that H₂O₂ produces oxidative stress in endothelial cells by increasing intracellular O₂⁻ levels through NOS and NADPH oxidase. These findings suggest a complex interaction between H₂O₂ and oxidant-generating enzymes that may contribute to endothelial dysfunction.     

Thermostability of β-xylosidase from Aspergillus sydowii MG49

Heating of Aspergillus β-xylosidase at 85°C ± 1°C and pH 5.5–6.0 (optimum for activity), causes irreversible, covalent thermoinactivation of the enzyme, involving oxidation of the thiol groups that are required for catalysis. Exogenous addition of cysteine, DTT, GSH and mercaptoethanol stabilizes the enzyme by extending its half-life. A similar effect is also exhibited by bivalent cations like Mg²⁺, Mn²⁺, Co²⁺, Ca²⁺and Zn²⁺ while, on the other hand Cu²⁺ accelerates thermoinactivation. Chemical modification of crude β-xylosidase with cross-linking agents like glutaraldehyde or covalent immobilization to a nonspecific protein like gelatin and BSA also enhances enzyme thermostability. These results suggest that addition of thiols and bivalent metal ions to a crude β-xylosidase preparation or immobilization/chemical modification enhances its thermal stability, thus preventing loss of catalytic activity at elevated temperatures.     

Purification and characterization of an endocellulase from the thermophilic fungus Chaetomium thermophilum CT2

Chaetomium thermophilum CT2 produced endocellulases at 50 °C, when grown on 2% microcrystalline cellulose, 1% soluble starch, and 0.4% yeast extract medium. A major endocellulase component was purified to homogeneity by fractional ammonium sulphate precipitation, ion-exchange chromatography on DEAE-Sepharose, Phenyl-Sepharose hydrophobic interaction chromatography and gel filtration on Sephacryl S-100. The molecular weight of the enzyme was estimated to be 67.8 kDa and the enzyme was found to be a glycoprotein containing 18.9% carbohydrate. The K_m of the purified enzyme for carboxymethyl cellulose, sodium salt (CMC), was 4.6 mg ml⁻¹. The enzyme displayed highest activity towards CMC and significantly lower activities towards phosphoric acid swollen cellulose and filter paper. The activity was enhanced in the presence of Na⁺, K⁺ and Ca²⁺ but inhibited by Hg²⁺, Zn²⁺, Ag⁺, Mn²⁺, Ba²⁺, Fe²⁺, Cu²⁺, Mg²⁺ and NH₄⁺. Optimum activity was at 60 °C and pH 4.0. The enzyme was stable over 60 min incubation at 60 °C and half-life at 70, 80 and 90 °C was approximately 45, 24 and 7 min, respectively.     

Reactions of the ferri-ferrocytochrome-c system with superoxide/oxygen and CO2/CO2 studied by fast pulse radiolysis

The reduction of ferricytochrome c by O₂⁻ and CO₂⁻ was studied in the pH range 6.6–9.2 and Arrhenius as well as Eyring parameters were derived from the rate constants and their temperature dependence. Ionic effects on the rate indicate that the redox process proceeds through a multiply-positively charged interaction site on cytochrome c. It is shown that the reaction with O₂⁻ and correspondingly with O₂ of ferrocytochrome c) is by a factor of approx. 10³ slower than warranted by factors such as redox potential. Evidence is adduced to support the view that this slowness is connected with the role of water in the interaction between O₂⁻/O₂ and ferri-ferrocytochrome c in the positively charged interaction site on cytochrome c in which water molecules are specifically involved in maintaining the local structure of cytochrome c and participate in the process of electron equivalent transfer.     

The use of chemically modified and unmodified cassava waste for the removal of Cd, Cu and Zn ions from aqueous solution

The use of different chemically modified cassava waste biomass for the enhancement of the adsorption of three metal ions Cd, Cu and Zn from aqueous solution is reported in this paper. Treating with different concentrations of thioglycollic acid modified the cassava waste biomass.

The sorption rates of the three metals were 0.2303 min⁻¹ (Cd²⁺), 0.0051 min⁻¹ (Cu²⁺), 0.0040 min⁻¹ (Zn²⁺) and 0.109 min⁻¹ (Cd²⁺), 0.0069 min⁻¹ (Cu²⁺), 0.0367 min⁻¹ (Zn²⁺) for 0.5 and 1.00 M chemically modified levels, respectively. The adsorption rates were quite rapid and within 20–30 min of mixing, about 60–80% of these ions were removed from the solutions by the biomass and that chemically modifying the binding groups in the biomass enhanced its adsorption capacity towards the three metals. The results further showed that increased concentration of modifying reagent led to increased incorporation, or availability of more binding groups, in the biomass matrix, resulting in improved adsorptivity of the cassava waste biomass. The binding capacity study showed that the cassava waste, which is a serious environmental nuisance, due to foul odour released during decomposition, has the ability to adsorb trace metals from solutions.     

A nitrite reductase with cytochrome oxidase activity from Micrococcus denitrificans

The formation of nitrite reductase and cytochrome c in Micrococcus denitrificans was repressed by O₂. The purified nitrite reductase utilized reduced forms of cytochrome c, phenazine methosulphate, benzyl viologen and methyl viologen, respectively, as electron donors. The enzyme was inhibited by KCN, NaN₃ and NH₂OH each at 1 mM, whereas CO and bathocuproin, diethyl dithiocarbamate, o-phenanthroline and ,'-dipyridyl at 1 mM concentrations were relatively ineffective. The purified enzyme contains cytochromes, probably of the c and a₂ types, in one complex. A K_m of 46 μM for NO₂⁻ and a pH optimum of 6.7 were recorded for the enzyme. The molecular weight of the enzyme was estimated to be around 130000, and its anodic mobility was 6.8·10⁻⁶ cm²·sec⁻¹·V⁻¹ at pH 4.55.

The most highly purified nitrite reductase still exhibited cytochrome c oxidase activity with a K_m of 27 μM for O₂. This activity was also inhibited by KCN, NaN₃ and NH₂OH and by NO₂⁻.

A constitutive cytochrome oxidase associated with membranes was also isolated from cells grown anaerobically with NO₂⁻. It was inhibited by smaller amounts of KCN, NaN₃ and NH₂OH than the cytochrome oxidase activity of the nitrite reductase enzyme and also differed in having a pH optimum of about 8 and a K_m for O₂ of less than 0.1 μM. Spectroscopically, cytochromes b and c were found to be associated with the constitutive oxidase in the particulate preparation. Its activity was also inhibited by NO₂⁻.

The physiological role of the cytochrome oxidase activity associated with the purified nitrite reductase is likely to be of secondary importance for the following reasons: (a) it accounts for less than 10% of total cytochrome c oxidase activity of cell extracts; (b) the constitutive cytochrome c oxidase has a smaller K_m for O₂ and would therefore be expected to function more efficiently especially at low concentrations of O₂.     

Effect of chloride ions on the S-state distribution and deactivation kinetics in preparations of the cyanobacterium Anacystis nidulans

The roles of Ca²⁺ and Cl⁻ on the photosynthetic O₂ yield under flash illumination have been examined in EDTA-washed preparations of the cyanobacterium Anacystis nidulans. Especially the effect of Cl⁻ deficiency on the O₂ yield and on the S-state distribution was analyzed. As the results show, omission of both Ca²⁺ and Cl⁻ (Mn²⁺ present) almost totally inhibited O₂ evolution. When Ca²⁺ was replaced by Na⁺, a substantial reduction of the O₂ yield was observed, but only a minor change in the S-state distribution occurred. However, when Cl⁻ was displaced by NO⁻₃, which is equivalent to Cl⁻ deficiency of the water-splitting complex, a substantial reduction of the O₂ yield and in addition a significant change in the S-state distribution was observed. The comparison of deactivation kinetics in NO⁻₃ containing samples with those in control samples indicated that Cl⁻ deficiency allowed accumulation of oxidizing equivalents up to the S₃ state but modified the final step of O₂ evolution. Moreover, those centers which advanced to the S₃ state in the absence of Cl⁻ deactivated in a special way which involved a faster deactivation of S₂ and an increased formation of S₋₁.     

Evidence that de novo protein synthesis participates in a time-dependent augmentation of the chemotactic peptide-induced respiratory burst in neutrophils: Effects of recombinant human colony stimulating factors and dihydrocytochalasin B

We examined the effects of the recombinant human colony stimulating factors GM-CSF and G-CSF, cycloheximide (a protein synthesis inhibitor) and dihydrocytochalasin B (a microfilament disrupting agent) upon FMLP (N-formyl-methionyl-leucylphenylalanine)-stimulated O₂ ⁻ production by neutrophils. We confirmed a time dependent augmentation of O₂ ⁻ production following preincubation of neutrophils either alone or with colony stimulating factors. Furthermore, we found that GM-CSF, but not G-CSF, increased O₂ ⁻ production at some concentrations of the stimulus. Preincubation of neutrophils with cycloheximide in the absence of CSF caused a marked fall in O₂⁻-production that was first evident at 2 hours. The fall in O₂⁻-forming capacity caused by cycloheximide was much less pronounced if dihydrocytochalasin B was also included in the preincubation buffer. These findings suggest a previously unrecognized role for de novo protein synthesis in maintaining the ability of neutrophils to manufacture O₂⁻, and support earlier studies indicating that the cycling of FMLP receptors between the cell membrane and an intracellular compartment is important in determining the magnitude of the respiratory burst in FMLP-stimulated neutrophils.     

Characterization of the particulate nitrite oxidase and its component activities from the chemoautotroph Nitrobacter agilis

1. Difference spectra, at room and liquid N₂ temperatures, of S₂O₄²⁻-, and NO₂⁻-reduced intact cells and cell-free preparations of Nitrobacter agilis demonstrated the presence of cytochromes of the c- and a-types. Reduction of cytochromes by succinate, and to a limited extent, by NADPH also occurred, provided KCN (0.1 mM) was also present.

2. A particulate, heat-labile nitrite oxidase having an absolute requirement for O₂ was prepared from N. agilis cells using sonic oscillation and differential centrifugation. The particles also possessed NADH oxidase, succinoxidase, formate oxidase and traces of NADPH oxidase activity. The stoichiometry of the nitrite oxidase reaction approached the theoretical value of 2 moles of NO₂⁻ consumed per mole of O₂ consumed. The pH optimum of the nitrite oxidase system shifted to progressively more alkaline values as the NO₂⁻ concentration was increased, changing from a pH value of 6.8 at 0.6 mM KNO₂ to pH 8.0 at 0.01 M KNO₂ with apparent K_m's of 0.2 and 1.2 mM NO₂⁻, respectively. Computations of the HNO₂ concentrations present under the above conditions showed an approx. 500-fold greater affinity for HNO₂ which was independent of pH, suggesting the involvement of HNO₂ as both a substrate and an inhibitor (at higher concentrations) of the nitrite oxidase system. The marked inhibition by NaN₃, NaCN and Na₂S, as well the light-reversible inhibition by CO, indicated the presence of cytochrome oxidase which was subsequently characterized. NO₂⁻ proved to be a competitive inhibitor of the nitrite oxidase system.

3. The particulate preparation also possessed a heat-labile nitrite-cytochrome c reductase activity which was energy independent and routinely measured under anaerobic conditions. As in the case of nitrite oxidase, the affinity of the enzyme for NO₃⁻ increased as the pH was lowered, but the pH optimum remained unaffected. In terms of calculated HNO₂ concentration an approximately constant K_m of about 0.2 μM was estimated at the several pH's examined. The inhibition by NO₃⁻ was shown to be competitive. The marked sensitivity of the reductase to several metal-binding agents implicated a metal component in the electron transport chain at the site prior to cytochrome c.

4. The membrane-like composition of the nitrite oxidase system is indicated.     

食品添加剂和金属离子对高粱泡红色素稳定性的影响

探讨了5种常用食品添加剂和7种金属离子对高粱泡红色素稳定性的影响。结果表明:葡萄糖、蔗糖和苯甲酸钠对高粱泡红色素稳定性无不良影响,其中葡萄糖、蔗糖有不同程度的增(护)色效果;柠檬酸能显著提高色素的稳定性;而VitC能促进色素的氧化降解,有明显的破坏作用。金属离子中,Na⁺、Mg²⁺、Al³⁺、Zn²⁺等对高粱泡红色素稳定性无影响,且有一定增色作用;较高浓度(≥0.0025mol/L)Mn²⁺有一定不良影响,而Cu²⁺、Fe³⁺则有明显的破坏作用。     

Oxidative phosphorylation coupled to oxygen uptake and nitrate reduction in Micrococcus denitrificans

A procedure is described for preparing particles from cells of Micrococcus denitrificans which were broken osmotically after treatment with lysozyme.

1. 1. The preparations catalysed ATP synthesis coupled to O₂ uptake or NO₃⁻ reduction. With NADH or succinate as the electron donors the P:O ratios were about 1.5 and 0.5, respectively; and the P:NO₃⁻ ratios were about 0.9 and 0.06, respectively.

2. 2. Addition of ADP or P_i to the reaction mixture increased the rates of NADH-dependent O₂ uptake and NO₃⁻ reduction. Addition of 1 mM 2,4-dinitrophenol, which inhibited phosphorylation by 50–60%, increased the basal rates of electron transport.

3. 3. Evidence derived from spectrophotometry and from the differential inhibition by antimycin A of O₂ and NO₃⁻ reduction leads to the conclusion that the nitrate reductase interacted with the respiratory chain in the region of the b-type cytochrome, and that the c-type cytochrome present was not involved in the reduction of NO₃⁻ to NO₂⁻.