Application of the theory of oxygen transfer: determination of the Michaelis constant of glucose oxidase with respect to oxygen]

The oxidation of beta-D-glucose with glucose oxidase generally requires oxygen, which, under normal conditions is present at low concentrations in the reaction medium. Experiments show that glucose oxidase is no longer saturated by oxygen at enzyme concentrations greater than 0.4 mg.ml1. This is due to the decrease in the oxygen concentration of the solution. The value of the oxygen mass transfer coefficients and dissolved oxygen concentrations are determined. These dissolved oxygen concentrations are found to correlate with direct measurements with an oxygen electrode. From this, the Michaelis constant of glucose oxidase for oxygen is calculated. These experiments also show that oxygen is a limiting factor for this reaction.     

Comparison of polarographic and chemical measurements of oxygen uptake in complex media: the example of lipoxygenase reaction.

A polarographic method using a Clark oxygen electrode was used to assess oxygen concentrations in model and complex media before and during the lipoxygenase-catalyzed oxidation of linoleic acid to hydroperoxides. The results were in good agreement with those obtained with the chemical determination of dissolved oxygen. The electrode correctly responded when the dissolved oxygen concentration was decreased by the addition of water activity depressors (sorbitol, sucrose, glucose). The influence of the medium components on the gas solubility was discussed. The linear relationship between partial pressure of oxygen (determined by polarographic method) and the oxygen concentration (determined by chemical method) indicated that the Clark oxygen electrode can be used to study enzyme reactions consuming or evolving oxygen in non-Newtonian media.     

The effect of myoglobin-facilitated oxygen transport on the basal metabolism of papillary muscle.

A mathematical model of oxygen diffusion into cylindrical papillary muscles is presented. The model partitions total oxygen flux into its simple and myoglobin-facilitated components. The model includes variable sigmoidal, exponential, or hyperbolic functions relating oxygen partial pressure to both fractional myoglobin saturation and rate of oxygen consumption. The behavior of the model was explored for a variety of saturation- and consumption-concentration relations. Facilitation of oxygen transport by myoglobin was considerable as indexed both by the elevation of oxygen partial pressure on the longitudinal axis of the muscle and by the fraction of total oxygen flux at the muscle center contributed by oxymyoglobin. Despite its facilitation of oxygen flux at the muscle center, myoglobin made only a negligible contribution to the total oxygen consumption averaged over the muscle cross-section. Hence the presence of myoglobin fails to explain either the experimentally determined basal metabolism-muscle radius relation or the stretch effect observed in isolated papillary muscle.     

Measurement of low levels of oxygen and their effect on respiration in cell suspensions maintained in an open system

The presence of low levels of oxygen may have profound effects on the cytotoxic activity of radiation, radiosensitizers, and bioreductive alkylating agents. As others have shown, low oxygen tensions may significantly alter rates of cellular and chemical oxygen consumption. When experiments are performed at very low oxygen concentrations, the opposing effects of oxygen leakage into and cellular/chemical oxygen consumption from the system can lead to unpredictable results. Use of a newly designed, highly sensitive Clark-type oxygen sensor has permitted accurate and reproducible measurement of low levels of oxygen. Cellular depletion of oxygen at various cell densities has been monitored for a series of oxygen tensions in solution and the corresponding respiration rates have been calculated. Although oxygen depletion was found to be quite significant at low oxygen tensions, not all oxygen present could be removed by cellular respiration. Respiration rate decreased as oxygen tension decreased and approached zero at low oxygen tensions. This result was independent of cell density. A model is presented to account for the observed effect of oxygen tension on cellular oxygen utilization.     

Oxygen Evolution and Oxygen Uptake by Isolated Chloroplasts of Wheat Irradiated with Monochromatic Light, without the Addition of an Oxidant

The oxygen exchange obtained when isolated chloroplasts of wheat are irradiated, without the addition of a Hill oxidant, has been investigated. Depending on the wavelength, two types of oxygen exchange are obtained. In light absorbed by both photosystems an oxygen gush appears directly upon irradiation. This oxygen evolving reaction is soon replaced by an oxygen uptake which is present until the end of the irradiation period. In light absorbed mainly in photosystem I, no oxygen gush can be observed, instead an oxygen uptake appears directly upon irradiation. An oxygen evolving process can also be observed in irradiations performed with photo-system I light, but this process appears after 10–15 seconds of irradiation. The influence of various external factors on the oxygen gush and the oxygen uptake, e.g. different wavelengths, light intensity, length of the dark periods between irradiations, was studied. The results show that the oxygen evolving reaction appearing upon irradiation with light absorbed by photosystem II and I, reflect the reduction of an oxidant, probably plasto-quinone, in the electron transport chain between the two photosystems. The reoxidation of this oxidant can be brought about after irradiating with light absorbed in photosystem I, or by prolonging the dark period between irradiations, or through some unknown process connected to photosystem II. The oxygen uptake which consists of two components, one appearing directly upon irradiation and the other one appearing after about 10 seconds of irradiation, confirms earlier observations that oxygen can be reduced in photosystem I. The electrons for the oxygen uptake appearing directly upon irradiation, are obtained from the reduced intermediates in the electron transport chain between the two photosystems. The electrons for the other oxygen uptake process are obtained from a reductant in the chloroplasts with access to the carrier chain between the photosystems. Whether the two oxygen uptake reactions reflect two sites of interaction of oxygen with the electron transport chain or only one site is discussed.     

The steady-state transport of oxygen through hemoglobin solutions

The steady-state transport of oxygen through hemoglobin solutions was studied to identify the mechanism of the diffusion augmentation observed at low oxygen tensions. A novel technique employing a platinum-silver oxygen electrode was developed to measure the effective diffusion coefficient of oxygen in steady-state transport. The measurements were made over a wider range of hemoglobin and oxygen concentrations than previously reported. Values of the Brownian motion diffusion coefficient of oxygen in hemoglobin solution were obtained as well as measurements of facilitated transport at low oxygen tensions. Transport rates up to ten times greater than ordinary diffusion rates were found. Predictions of oxygen flux were made assuming that the oxyhemoglobin transport coefficient was equal to the Brownian motion diffusivity which was measured in a separate set of experiments. The close correlation between prediction and experiment indicates that the diffusion of oxyhemoglobin is the mechanism by which steady-state oxygen transport is facilitated.     

Effects of Some Chemicals on the Oxygen Evolution and Oxygen Uptake in Isolated Wheat Chloroplasts Irradiated without the Addition of an Oxidant

The oxygen exchange, obtained when isolated chloroplasts of Triticum aestivum, wheat, are irradiated without the addition of a Hill oxidant has been investigated using an oxygen electrode. Ascorbate, catalase, 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone(DBMIB), diethyldithio-carbamate (DEDT), dichlorophenylmethylurea (DCMU), and potassium cyanide were added to the Chloroplasts in order to investigate the oxygen exchange. At least two oxygen uptake reactions, one sensitive to catalase and one catalase-insensitive, appeared upon irradiation. Hydrogen peroxide was the product of the oxygen uptake in the former process, and water was the reductant. The formation of hydrogen peroxide was probably associated with photosystem I. The other oxygen consuming reaction was found to be insensitive to both catalase and potassium cyanide. After the chloroplasts had been treated with DCMU, it was possible to show that the catalase-insensitive oxygen uptake was localized in photosystem I, and that a cyclic electron transport system or some endogenous reductant (-s) acted in the oxygen uptake. Addition of ascorbate or DEDT to the chloroplasts led to an enhanced oxygen uptake in 710 nm light. This was probably due to the effect of these compounds on the superoxide radical ion formed in photosystem I. The stimulated oxygen uptake was only weakly affected by catalase, indicating that hydrogen peroxide was not a product of this oxygen uptake. Addition of DEDT and potassium cyanide inhibited (strongly respectively weakly) the oxygen uptake when photosystem II was functioning. The effect of these compounds was probably due to an inhibition of the electron transport at the plastocyanin. DBMIB inhibited the oxygen uptake reactions and the cooperation between the two photosystems. The cooperation between the photosystems was also studied in DCMU-treated chloroplasts. The reactions in photosystem II, measured as oxygen evolution, were more inhibited than the coupling between the photosystems. The oxygen “gush” appearing upon irradiation in light of 650 nm was not affected by a DBMIB-treatment, showing that the oxygen evolution was due to the reduction of plastoquinone. The reoxidation in the dark of the plastoquinone pool was stimulated by DBMIB and potassium cyanide indicating that an oxygen uptake could be associated with plastoquinone. The sites of interaction of oxygen with the electron transport pathways in chloroplasts, and the different reductants for the oxygen consuming reactions are discussed.     

A model study of intracellular oxygen gradients in a myoglobin-containing skeletal muscle fiber.

A theoretical two-dimensional model is used to investigate oxygen gradients in a red skeletal muscle fiber. The model describes the steady state, free and myoglobin-facilitated diffusion of oxygen into a respiring cylindrical muscle fiber cross section. The oxygen tension at the sarcolemma is assumed to vary along the sarcolemma as an approximation to the discrete capillary oxygen supply around the fiber. Maximal oxygen gradients are studied by considering parameters relevant to a maximally-respiring red muscle fiber. The model predicts that angular variations in the oxygen tension imposed at the sarcolemma due to the discrete capillary sources do not penetrate deeply into the fiber over a range of physiological values for myoglobin concentration, diffusion coefficients, number of surrounding capillaries, and oxygen tension level at the sarcolemma. Also, the oxygen tension in the core of the fiber is determined by the average oxygen tension at the sarcolemma. The drop in oxygen tension from fiber periphery to core, however, does depend significantly on the myoglobin concentration, the oxygen tension level at the sarcolemma, and the oxygen and myoglobin diffusivities. This dependence is summarized by calculating the minimum average sarcolemmal oxygen tension for maximal respiration without the development of an intracellular anoxic region. For a myoglobin-rich muscle fiber (0.5 mM myoglobin), the model predicts that maximal oxygen consumption can proceed with a relatively flat (less than 5 mm Hg) oxygen tension drop from fiber periphery to core over a large range for diffusion coefficients.     

The catalytic effect of the carcinogen "4-nitroquinoline-N-oxide" on the oxidation of vitamin C.

The carcinogen 4-nitroquinoline-N-oxide was found to mediate the reaction between ascorbate and oxygen. The oxidation of ascorbate was initiated by the production of the nitro radical anion which reacted with oxygen to produce the oxygen superoxide radical anion, peroxide and hydroxyl radical. The production of partially reduced oxygen intermediates resulted in additional reactions with ascorbate. The consumption of oxygen could be either completely blocked by reacting the nitro radical with ferricytochrome c or partially blocked by the combined effects of superoxide dismutase and catalase. The consumption of oxygen could be enhanced by reducing the hydroxyl radicals with dimethylsulfoxide.     

Oxygen regulation of nitrate uptake in denitrifying Pseudomonas aeruginosa

Oxygen had an immediate and reversible inhibitory effect on nitrate respiration by denitrifying cultures of Pseudomonas aeruginosa. Inhibition of nitrate utilization by oxygen appeared to be at the level of nitrate uptake, since nitrate reduction to nitrite in cell extracts was not affected by oxygen. The degree of oxygen inhibition was dependent on the concentration of oxygen, and increasing nitrate concentrations could not overcome the inhibition. The inhibitory effect of oxygen was maximal at approximately 0.2% oxygen saturation. The inhibition appeared to be specific for nitrate uptake. Nitrite uptake was not affected by these low levels of aeration, and nitrite reduction was only partially inhibited in the presence of oxygen. The regulation of nitrate respiration at the level of transport by oxygen may represent a major mechanism by which the entire denitrification pathway is regulated in P. aeruginosa.     

High tolerance of methanogens in granular sludge to oxygen

This research assessed the effect of oxygen exposure on the methanogenic activity of anaerobic granular sludges. The toxicity of oxygen to acetoclastic methanogens in five different anaerobic granular sludges was determined in serum flasks with effective gas-to-liquid volumes of 4.65 to 1. The amount of oxygen that caused 50% inhibition of the methanogenic activity after 3 days of exposure ranged from 7% to 41% oxygen in the head space. These results indicate that methanogens located in granular sludge have a high tolerance for oxygen. The most important factor contributing to the tolerance was the oxygen consumption by facultative bacteria metabolizing biodegradable substrates. Uptake of oxygen by these bacteria creates anaerobic microenvironments where the methanogenic bacteria are protected. The results also indicate that methanogens in sludge consortia still have some tolerance to oxygen, even in the absence of facultative substrate for oxygen respiration. (c) 1993 John Wiley & Sons, Inc.     

Oxygen regulation of nitrate uptake in denitrifying Pseudomonas aeruginosa.

Oxygen had an immediate and reversible inhibitory effect on nitrate respiration by denitrifying cultures of Pseudomonas aeruginosa. Inhibition of nitrate utilization by oxygen appeared to be at the level of nitrate uptake, since nitrate reduction to nitrite in cell extracts was not affected by oxygen. The degree of oxygen inhibition was dependent on the concentration of oxygen, and increasing nitrate concentrations could not overcome the inhibition. The inhibitory effect of oxygen was maximal at approximately 0.2% oxygen saturation. The inhibition appeared to be specific for nitrate uptake. Nitrite uptake was not affected by these low levels of aeration, and nitrite reduction was only partially inhibited in the presence of oxygen. The regulation of nitrate respiration at the level of transport by oxygen may represent a major mechanism by which the entire denitrification pathway is regulated in P. aeruginosa.     

Uptake of oxygen,release and degradation of hydrogen peroxide by Streptococcus mutans NCTC 10449

Streptococcus mutans NCTC 10499 was cultured under glucose limitation in a chemostat at varying oxygen supply. The rates of oxygen uptake and hydrogen peroxide degradation by cells from the cultures were measured polarographically using a Clark electrode. Oxygenation of the chemostat culture led to adaptation of the organism to oxygen, in that the maximum oxygen uptake rate of the cells was higher when the cells were grown at higher rate of oxygen supply. It is noted that anaerobically grown cells still exhibited significant oxygen uptake. The rate of oxygen uptake followed saturation-type kinetics and Ks values of cells for oxygen were in the micromole range. Hydrogen peroxide accumulation was not observed in aerated chemostat cultures. However, anaerobically grown cells accumulated H2O2 when exposed to oxygen. Cells from aerated cultures did not accumulate hydrogen peroxide. This may be explained by the fact that the rate of hydrogen peroxide degradation was consistently higher than the rate of oxygen uptake.     

Aroma Components of Fresh Sugar Cane Juice

To study the behavior of Bifidobacterium toward oxygen, oxygen uptake was investigated in detail. The cells of Bifidobacterial strains absorbed considerable amounts of oxygen. The exogenous oxygen uptake activity changed depending upon the period of incubation. Bifidobacterial cells also had high endogenous oxygen uptake, which was, in B. longum strains, as high as about 80% of the exogenous oxygen uptake activity. Bifidobacterial cells accumulated considerable amounts of polysaccharide, which was associated with cellular growth. By incubating the cultivated cells in a glucose-free medium, the endogenous oxygen uptake activity was decreased with a decrease of intracellular polysaccharide. Therefore it was postulated that the high endogenous oxygen uptake activity of Bifidobacterium was owing to the metabolism of intracellular polysaccharide. The enzymatic activity, which was involved in the mechanism of oxygen uptake, was also investigated.     

The relationship between critical oxygen level and antibiotic synthesis of capreomycin and cephalosporin C

This paper illustrates the type of conclusions which may be reached by the use of an electrode for measuring oxygen tension in fermentation broths. The critical oxygen level, based on oxygen uptake, and the minimum oxygen tension necessary for maximum antibiotic synthesis for two commercial antibiotic fermentations, capreomycin and cephalosporin C, were studied. Capreomycin yields were depressed at an oxygen tension which was slightly below the critical oxygen level. Cephalosporin C synthesis was depressed at an oxygen tension higher than the critical oxygen level.     

The functional prognosis of physical working ability under conditions of extreme high altitude

The physical performance of climbers, those making high-altitude ascensions up to 8000 m, without additional oxygen was measured. Some functional criteria of the organism adaptation to exhausting physical loading at the high altitudes were selected. It was established that the forecasting of a successful ascension could be improved if the potential maximal oxygen uptake was added to the standard definition parameters of aerobic metabolism such as the maximal oxygen uptake and anaerobic threshold of oxygen uptake. The potential maximal oxygen uptake is considered to be the parameter of maximal oxygen uptake reserve growing in condition of realization of adaptive reaction to hypoxia.     

Growth of Trichosporon cutaneum under oxygen limitation: kinetics of oxygen uptake

The relationship between oxygen concentration and growth rate in the yeast Trichosporon cutaneum was studied. In order to establish the conditions for purely oxygen-limited growth, the cells were first grown in a carbon-limited chemostat, and kinetic parameters determined. The cells were then grown in an oxygen-limited chemostat at different dilution rates yielding different oxygen uptake rates. The steady-state dissolved oxygen tension was found at each dilution rate and the corresponding equilibrium dissolved oxygen tension was found at each dilution rate and the corresponding equilibrium dissolved oxygen concentration determined in the effluent medium. The relationship between oxygen concentration and growth rate followed Monod-type kinetics with an apparent K(O) of 4.38 x 10(-6)M.     

Geological constraints on the origin of oxygenic photosynthesis

This article examines the geological evidence for the rise of atmospheric oxygen and the origin of oxygenic photosynthesis. The evidence for the rise of atmospheric oxygen places a minimum time constraint before which oxygenic photosynthesis must have developed, and was subsequently established as the primary control on the atmospheric oxygen level. The geological evidence places the global rise of atmospheric oxygen, termed the Great Oxidation Event (GOE), between ~2.45 and ~2.32 Ga, and it is captured within the Duitschland Formation, which shows a transition from mass-independent to mass-dependent sulfur isotope fractionation. The rise of atmospheric oxygen during this interval is closely associated with a number of environmental changes, such as glaciations and intense continental weathering, and led to dramatic changes in the oxidation state of the ocean and the seawater inventory of transition elements. There are other features of the geologic record predating the GOE by as much as 200–300 million years, perhaps extending as far back as the Mesoarchean–Neoarchean boundary at 2.8 Ga, that suggest the presence of low level, transient or local, oxygenation. If verified, these features would not only imply an earlier origin for oxygenic photosynthesis, but also require a mechanism to decouple oxygen production from oxidation of Earth’s surface environments. Most hypotheses for the GOE suggest that oxygen production by oxygenic photosynthesis is a precondition for the rise of oxygen, but that a synchronous change in atmospheric oxygen level is not required by the onset of this oxygen source. The potential lag-time in the response of Earth surface environments is related to the way that oxygen sinks, such as reduced Fe and sulfur compounds, respond to oxygen production. Changes in oxygen level imply an imbalance in the sources and sinks for oxygen. Changes in the cycling of oxygen have occurred at various times before and after the GOE, and do not appear to require corresponding changes in the intensity of oxygenic photosynthesis. The available geological constraints for these changes do not, however, disallow a direct role for this metabolism. The geological evidence for early oxygen and hypotheses for the controls on oxygen level are the basis for the interpretation of photosynthetic oxygen production as examined in this review.     

Studies on reactions of illuminated chloroplasts. III. Simultaneous photoproduction and consumption of oxygen studied with oxygen isotopes

The evolution of oxygen, as in a typical Hill reaction, during periods of net oxygen consumption by illuminated chloroplasts has been demonstrated through kinetic studies with oxygen isotopes. The ability of molecular oxygen to function as a Hill oxidant is thus confirmed.