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[Effects of technological additives and heating range on some chemical and physical changes in canned meat. 2. Changes in redox potentials and selected quality characteristics].

The influence of technological additives and the range of heating on the redox potential, as well as on some quality features of canned meat was examined. The experiments showed, that the time of storage and the degree of heating of model preserves of meat influence on the redox potential. The technological additions as polyphosphates, ascorbic acid, gelatine and mixtures of these substances influence less on the redox potential. The analysis of each experimental factor showed, that on the secretion of meat juice occurring during can pasteurization or sterilization influence all experimental factors, as the kind of heating, the time of storage as well as the kind and the quantity of technological additives. The highest secretion of meat juice was found in cans with addition of ascorbic acid. Cans with addition of gelatine had the smallest content of jelly and consequently the lowest secretion of meat juice. It was also found a certain relation between the level of redox potential and the tested quality features of the model meat preserves.

Ascorbic Acid↗

A field study of some swimming-pool waters with regard to bacteria, available chlorine and redox potential.

The waters of one indoor bath and three outdoor baths were examined once an hour during 3 days (bath 1) or 6 days, for available chlorine, redox potential, permanganate number, ammonium, nitrate and total nitrogen, total bacterial count at 22 degrees C., total bacterial count at 37 degrees C. and faecal coliform bacteria. The weather, number of swimmers and the chlorine gas addition were continuously registered, and the pH was checked a few times at each bath. In bath 1, an indoor pool with aluminium sulphate precipitation about once a week and with sand filters back-washed every 2 days, less than 10 bacteria/ml. were found in all samples. In bath 2, an outdoor pool with aluminium sulphate precipitation twice a week and with sand filters back-washed twice a week, also few bacteria were found. In bath 3, an outdoor pool with only filtering through sand filters back-washed about every 14 days, high bacterial counts were found every day except the first, when the filters had been newly back-washed. In bath 4, an outdoor pool with only filtering through sand filters back-washed about once a week, high bacterial counts were found now and then during the first 4 days when the weather was warm, but few bacteria were found the last 2 days when the weather was cold and windy, and there were few swimmers.Values from different analyses on the same sample showed relatively good correlation between the redox potential and the free available chlorine. In bath 3 both the redox potential and the available chlorine were weakly correlated to the bacterial count, but in bath 4 there was no such correlation. No other factors were well correlated with the bacterial count either.The bacterial counts at 22 degrees and 37 degrees C. were of the same order. No faecal coliforms were ever found. Use of these bacteria as indicator organism in swimming pools is criticized.The method of using certain minimum values of the free available chlorine as guarantee for a satisfactory bacteriological quality of the swimming pool water is also questioned. The degree of purity of the water is fundamentally connected with the disinfecting power of the available chlorine.Use of certain minimum values of the redox potential, according to these investigations, seems to be a method of somewhat greater accuracy. Provided that the methods of precipitation are performed correctly and filters are being back-washed often enough, then an automatically registering redox potential device, perhaps connected to the chlorine gas pump, ought to constitute a good control of the hygienic quality of a swimming-pool water. This must, however, always be completed by bacteriological examinations, preferably made at high bathing load.

Bacteria↗

Correlation of redox potential with state variables in cultures under controlled dissolved oxygen concentration and pH

In batch cultures for L-ornithine production in which dissolved oxygen concentration and pH were closely controlled, time changes of redox potential were observed in connection with the profiles of cell, glucose, and ornithine concentrations. It was found that the redox potential profile had four different phases reflecting the physiological state of the culture and that it was closely related to cell concentration change. Effects of glucose and ornithine on the redox potential were identified in a separate series of experiments. On the basis of the experimental results, a correlation of redox potential to glucose, cell, and ornithine concentrations has been proposed. The proposed correlation can be used for on-line estimation of ornithine concentration from on-line data of redox potential, glucose concentration, and cell concentration.

Journal Article↗

Structural changes caused by site-directed mutagenesis of tyrosine-98 in Desulfovibrio vulgaris flavodoxin delineated by 1H and 15N NMR spectroscopy: implications for redox potential modulation.

Flavodoxins mediate electron transfer at low redox potential between the prosthetic groups of other proteins. Interactions between the protein and the flavin mononucleotide cofactor shift both the oxidized/semiquinone and semiquinone/hydroquinone redox potentials significantly from their free-in-solution values. In order to investigate the possible role that the tyrosine at position 98 plays in this process, we have used heteronuclear three-dimensional NMR spectroscopy to determine the solution conformation of wild-type and four position-98 mutants, Y98W, Y98H, Y98A, and Y98R, of Desulfovibrio vulgaris flavodoxin. Assigned 1H and 15N resonances indicate that the secondary structure and topology of the proteins are identical. However, residues that undergo substantial mutation-induced changes in chemical shift are spread throughout the flavin cofactor binding site. Distance and dihedral angle constraints were used to generate solution structures for the wild-type and mutant proteins. Collectively, the mutant proteins have no gross conformational changes in the flavin binding site. The changes that do occur are minor and result from the different packing interactions required to accommodate the new side chain at position-98. The solvent accessibility and electrostatic nature of the flavin binding site in the mutant proteins are compared to those of the wild-type structure. The structural data support the hypothesis that the very low midpoint of the semiquinone/hydroquinone couple in the wild-type protein is modulated to a large extent by the energetically unfavorable formation of the flavin hydroquinone anion in the apolar environment of the flavin binding site.

Crystallography, X-Ray↗

Digestive proteinase activity in corn earworm (Helicoverpa zea) after molting and in response to lowered redox potential.

Insect digestive proteinases are often strongly influenced by ambient physicochemical conditions, such as pH, ionic strength, and oxidation-reduction potential. Although the effects of the former two parameters are well documented, the influence of redox potential on catalytic rates of digestive enzymes is not well understood. In this study, we manipulated the midgut redox potential of a generalist caterpillar (the corn earworm, Helicoverpa zea) by augmenting artificial diet with dithiothreitol, a powerful thiol reducing agent that lowers the redox potential in the lumen by 40-45 mV. Effects on total proteolytic activity, as well as on elastase, chymotrypsin, trypsin, leucine aminopeptidase, and carboxypeptidase A and B activities were measured using azocasein and nitroanilide model substrates. The profiles of proteinase activities in the epithelium and lumen were also monitored on days 1, 2, and 3 after the molt in penultimate instar larvae. Although the reducing agent strongly inhibited the activity of some proteinases in vitro, ingestion of the reducing diet failed to affect in vivo proteinase activities. There was also no effect on larval relative growth, consumption, or digestive efficiencies. We conclude that dietary reducing agents must lower midgut redox potential to below -40 mV to significantly impact digestive efficiency. Arch.

Animals↗

The protein moiety modulates the redox potential in cytochromes c.

Cytochrome c is one of the most thoroughly documented oxidoreduction proteins. Its electron transfer activity, which involves an association between the heme group and the polypeptidic chain, is correlated with the redox potential value of the heme group. The redox potential covers a wide range up to 0.8 V, an extreme case being observed in the low-potential cytochromes c from sulfate reducing bacteria. On of the main roles of the polypeptidic moiety consists of modulating the redox potential value of the heme group. In this paper, some structural factors that seem likely to be involved in maintaining the redox potential value are described.

Cytochrome c Group↗

Determination of the redox potential of deazariboflavin by equilibration with flavins.

The redox potential of deazariboflavin has been determined for pH values from 5.5 to 9.2 by equilibration with riboflavin and lumiflavin 3-acetate. The position of the equilibrium with riboflavin was measured spectrophotometrically and fluorimetrically; the equilibrium potential with lumiflavin 3-acetate was measured spectrophotometrically and potentiometrically. The Em7 for deazariboflavin was found to be--0.273 +/- 0.003 V against the standard hydrogen electrode. Equilibrium with flavodoxin at pH 9.5 and 10.0 was also used to determine the redox potential of deazariboflavin at high pH values. The pK of dihydrodeazariboflavin was found from the break in the potential vs. pH diagram and from spectrophotometric pH titration. The pK value obtained by both methods is 7.00 +/- 0.05. We found that borate, a product of the reducing agent borohydride, complexed with the ribityl sidechain of deazariboflavin, causing a shift in the pK for the reduced form to values of about 8.

Flavins↗

Cytoplasmic redox potential affects energetics and contractile reactivity of vascular smooth muscle.

Variations in the cytoplasmic redox potential (Eh) and NADH/NAD ratio as determined by the ratio of reduced to oxidized intracellular metabolite redox couples may affect mitochondrial energetics and alter the excitability and contractile reactivity of vascular smooth muscle. To test these hypotheses, the cytoplasmic redox state was experimentally manipulated by incubating porcine carotid artery strips in various substrates. The redox potentials of the metabolite couples [lactate]/[pyruvate]i and [glycerol 3-phosphate]/[dihydroxyacetone phosphate]i varied linearly (r=0.945), indicating equilibrium between the two cytoplasmic redox systems and with cytoplasmic NADH/NAD. Incubation in physiological salt solution (PSS) containing 10 mm pyruvate ([lact]/[pyr]=0.6) increased O2 consumption approximately 45% and produced anaplerosis of the tricarboxylic acid (TCA cycle), whereas incubation with 10 mm lactate-PSS ([lact]/[pyr]i=47) was without effect. A hyperpolarizing dose of external KCl (10 mM) produced a decrease in resting tone of muscles incubated in either glucose-PSS (-0.8+/-0.8 g) or pyruvate-PSS (-2.1+/-0.8 g), but increased contraction in lactate-PSS (1.5+/-0.7 g) (n=12-18, P<0.05). The rate and magnitude of contraction with 80 mm KCl (depolarizing) was decreased in lactate-PSS (P=0.001). Slopes of KCl concentration-response curves indicated pyruvate>glucose>lactate (P<0.0001); EC50 in lactate (29. 1+/-1.0 mM) was less than that in either glucose (32.1+/-0.9 mm) or pyruvate (32.2+/-1.0 mM), P<0.03. The results are consistent with an effect of the cytoplasmic redox potential to influence the excitability of the smooth muscle and to affect mitochondrial energetics.

Aerobiosis↗

Crystallographic investigation of the role of aspartate 95 in the modulation of the redox potentials of Desulfovibrio vulgaris flavodoxin.

The side chain of aspartate 95 in flavodoxin from Desulfovibrio vulgaris provides the closest negative charge to N(1) of the bound FMN in the protein. Site-directed mutagenesis was used to substitute alanine, asparagine, or glutamate for this amino acid to assess the effect of this charge on the semiquinone/hydroquinone redox potential (E(1)) of the FMN cofactor. The D95A mutation shifts the E(1) redox potential positively by 16 mV, while a negative shift of 23 mV occurs in the oxidized/semiquinone midpoint redox potential (E(2)). The crystal structures of the oxidized and semiquinone forms of this mutant are similar to the corresponding states of the wild-type protein. In contrast to the wild-type protein, a further change in structure occurs in the D95A mutant in the hydroquinone form. The side chain of Y98 flips into an energetically more favorable edge-to-face interaction with the bound FMN. Analysis of the structural changes in the D95A mutant, taking into account electrostatic interactions at the FMN binding site, suggests that the pi-pi electrostatic repulsions have only a minor contribution to the very low E(1) redox potential of the FMN cofactor when bound to apoflavodoxin. Substitution of D95 with glutamate causes only a slight perturbation of the two one-electron redox potentials of the FMN cofactor. The structure of the D95E mutant reveals a large movement of the 60-loop (residues 60-64) away from the flavin in the oxidized structure. Reduction of this mutant to the hydroquinone causes the conformation of the 60-loop to revert back to that occurring in the structures of the wild-type protein. The crystal structures of the D95E mutant imply that electrostatic repulsion between a carboxylate on the side chain at position 95 and the phenol ring of Y98 prevents rotation of the Y98 side chain to a more energetically favorable conformation as occurs in the D95A mutant. Replacement of D95 with asparagine has no effect on E(2) but causes E(1) to change by 45 mV. The D95N mutant failed to crystallize. The K(d) values of the protein FMN complex in all three oxidation-reduction states differ from those of the wild-type complexes. Molecular modeling showed that the conformational energy of the protein changes with the redox state, in qualitative agreement with the observed changes in K(d), and allowed the electrostatic interactions between the FMN and the surrounding groups on the protein to be quantified.

Alanine↗

Polyplex gene delivery modulated by redox potential gradients.

Polyplexes sensitive to redox potential gradients represent a promising class of vectors for delivery of nucleic acids. This review focuses on the recent advances in the development of these vectors. The biological rationale for the design of redox-sensitive polyplexes is discussed together with the basic synthetic approaches for introducing reducible disulfide bonds into the structure of the polyplexes. The biological properties of the redox-sensitive polyplexes of plasmid DNA, mRNA, antisense oligonucleotides and siRNA are reviewed with emphasis on in vitro cellular delivery, cytotoxicity and in vivo activity. Overall, redox-sensitive polyplexes represent a promising platform for further development as vectors for delivery of a wide variety of therapeutic nucleic acids.

Animals↗

The effect of the redox-potential on the retention of phosphorus in a small constructed wetland.

Building wetlands in small arable streams is a popular supplement to best management practice on arable fields. Particle bound phosphorus settles in the small constructed wetlands (CWs), receiving agricultural diffuse pollution. The sorption behavior of phosphorus is, however, redox-sensitive, and bound phosphorus may be remobilized in periods with low redox potential. This paper investigates changes in the redox potential in the free water of wetland Berg (Norway) during a three-year period, and how these redox changes affect the total phosphorus (TP) and total reactive phosphorus (TRP) retention. Despite eutrophic conditions in the wetland, the redox potential was never negative, and usually higher than 400 mV, indicating aerobic conditions. The relative retention was 440% and 43% for TP and TRP, respectively. The specific retention was 100 g TP and 43 g TRPm(-2) yr(-1). Loss of phosphorus was only observed during less than 19% of the total period of time. The net loss was less than 5% of the specific retention. The high positive redox potential probably conserves the redox-sensitive phosphorus in the wetland sediment as long as water fows through the CW.

Fresh Water↗

pH dependence of the redox potential of Pseudomonas aeruginosa cytochrome c-551.

The redox potential of Ps. aeruginosa cytochrome c-551 varies with pH between pH 5 and 8. The pH dependence can be analysed in terms of a pKa of 6.2 in the oxidised form and a pKa of 7.3 in the reduced form. The same pKa values are also observed in NMR spectra of the two oxidation states and the pKa of 7.3 is observed in titration of the visible absorption spectrum of the ferrocytochrome. From the NMR studies these pKa values have been assigned to the ionisation of one of the haem propionic acid groups. pH dependence of redox potential is of variable occurrence among cytochromes and the possible significance and basis of this variation is discussed.

Bacterial Proteins↗

Higher order structure contributes to specific differences in redox potential and electron transfer efficiency of root and leaf ferredoxins.

Plant type ferredoxin (Fd) is a small [2Fe-2S] cluster containing electron-transfer protein with a highly negative redox potential. Higher plants contain different iso-protein types of Fd in roots and leaves, reflecting the difference in redox cascades between these two tissues. We have combined subdomains of leaf and root Fds in recombinant chimeras, to examine structural effects and the relationship between groups of residues on redox potential, electron transfer, and protein-protein interactions. All chimeras had redox potentials that were intermediate to the wild type leaf and root Fds. Surprisingly, the largest differences resulted from exchange of the N-terminus, the region farthest from the redox center. Homology modeling and energy minimization calculations suggest that the N-terminal chimeras may indirectly influence redox potentials by structurally perturbing the active site. Measurements of electron transport and protein interaction indicate that synergistic interaction between the C- and N-terminal of root Fd bestows a specific high affinity for accepting electrons in the root type electron cascade, and that there is discrimination against photosynthetic electron donation to root Fd based on the C-terminus of the molecule. Taken together, the experimental and computational studies support a model in which higher order structure contributes to iso-protein specific interaction and electron-transfer properties.

Amino Acid Sequence↗

Direct measurement of the hydrogen-bonding effect on the intrinsic redox potentials of [4Fe-4S] cubane complexes.

To probe how H-bonding effects the redox potential changes in Fe-S proteins, we produced and studied a series of gaseous cubane-type analogue complexes, [Fe(4)S(4)(SEt)(3)(SC(n)H(2n+1))](2-) and [Fe(4)S(4)(SEt)(3)(SC(n)H(2n)OH)](2-) (n = 4, 6, 11; Et = C(2)H(5)). Intrinsic redox potentials for the [Fe(4)S(4)](2+/3+) redox couple involved in these complexes were measured by photoelectron spectroscopy. The oxidation energies from [Fe(4)S(4)(SEt)(3)(SC(n)H(2n)OH)](2-) to [Fe(4)S(4)(SEt)(3)(SC(n)H(2n)OH)](-) were determined directly from the photoelectron spectra to be approximately 130 meV higher than those for the corresponding [Fe(4)S(4)(SEt)(3)(SC(n)H(2n+1))](2-) systems, because of the OH...S hydrogen bond in the former. Preliminary Monte Carlo and density functional calculations showed that the H-bonding takes place between the -OH group and the S on the terminal ligand in [Fe(4)S(4)(SEt)(3)(SC(6)H(12)OH)](2-). The current data provide a direct experimental measure of a net H-bonding effect on the redox potential of [Fe(4)S(4)] clusters without the perturbation of other environmental effects.

Binding Sites↗

Antimalarial quinones: redox potential dependence of methemoglobin formation and heme release in erythrocytes.

A number of quinones have been shown to be effective antimalarials. In addition some have been shown to have oxidant effects on glucose-6-phosphate dehydrogenase-deficient red cells. To clarify the mechanism of this oxidant effect and to determine whether it is a general property of antimalarial quinones, the effects of five compounds on red cells were studied. Two, 5-hydroxy-6-demethyl pentaquine (5H6DP) and menadione, caused marked methemoglobin production and the insertion of heme into the red-cell membrane. The other three, atovaquone, daphnetin, and menoctone, did not. The insertion of heme into membranes caused diminished deformability, and this might account for the hemolytic effects. Redox potentials of all five compounds were measured. The two quinones that caused heme release, menadione and 5H6DP, had redox potentials of -141 +/- 12 and -97 +/- 25 mV, respectively, which are similar to the reported redox potentials for hemoglobin. The other three quinones had redox potentials that were either significantly higher or lower. Thus only quinones of the appropriate redox potentials are likely to be toxic to red cells.

Antimalarials↗

pH-dependent redox potential: how to use it correctly in the activation energy analysis.

The activation barrier (the activation free energy) for the reaction's elementary act proper does not depend on the presence of reactants outside the reaction complex. The barrier is determined directly by the concentration-independent configurational free energy. In the case of redox reactants with pH-dependent redox potential, only the pH-independent quantity, the configurational redox potential enters immediately into expression for activation energy. Some typical cases of such reactions have been discussed (e.g., simultaneous proton and electron detachment, acid dissociation followed by oxidation, dissociation after oxidation, and others). For these mechanisms, the algorithms for calculation of the configurational redox potential from the experimentally determined redox potentials have been described both for the data related to a dissolved reactant or to a prosthetic group of an enzyme. Some examples of pH-dependent enzymatic redox reactions, in particular for the Rieske iron-sulfur protein, have been discussed.

Electron Transport Complex III↗

Regulation of the flavin redox potential by flavin-binding antibodies.

Single-chain Fv antibody fragments binding different flavin forms [10-(5'-carboxybutyl-)flavin (Fl[ox]) and 10-(5'-carboxybutyl)-1,5-dihydroflavin (Fl[red])] have been generated from an antibody phage-display library to study how a protein environment regulates the redox potential, starting from a protein other than a natural flavoprotein. These 'flavobodies' are characterized by time-resolved and steady-state fluorescence spectroscopy, by competitive ELISA methods (mapping of the antigen-binding site), and by molecular modelling. The three-dimensional models of the antigen-binding sites are consistent with the experimental results. Binding of anti-Fl(red) 5 to flavin increases the redox potential, mainly due to an Arg residue interacting with the flavin N1. Thus anti-Fl(red) 5 shows an 'oxidase-like' redox-potential behaviour, confirming the idea that positively charged residues in the vicinity of N1 increase the redox potential. The results obtained with anti-Fl(ox), which do not resemble a natural flavoprotein, show that when the pyrimidine-like nucleus of the flavin is not involved in binding, the redox potential is not significantly affected. These results are in contrast to those obtained with chicken riboflavin-binding protein.

Amino Acid Sequence↗

Redox potential and survival of virulent Treponema pallidum under microaerophilic conditions.

A strongly reduced culture medium, capable of maintaining the virulence of Treponema pallidum (Nichols) for several days, was exposed to an atmosphere of 3% oxygen in nitrogen for 2-3 days before inoculation with T pallidum. By using various volumes of medium in uniform tubes a range of redox potentials (Ecal) from -94 mV to -325 mV was produced depending on the surface area-to-volume ratios of the medium. The anaerobic medium had an Ecal value of -387 mV. The medium was inoculated with T pallidum and incubated in an atmosphere of 3% oxygen. The survival of treponemes at different redox potentials was monitored by observing the retention of motility and by measuring the latent period of infection after inoculation of the cultures into the shaved backs of rabbits. Under these conditions T pallidum survived longest at low (electronegative) redox potential. An inverse linear relationship was observed between the redox potential of the culture medium and the survival of T pallidum, as measured by the time required for a 90% reduction of virulent organisms. No optimum redox potential was detected, the most electronegative medium (-325 mV, Ecal) giving the best survival.

Bacteriological Techniques↗