Simultaneous Measurement Of Oxygen Consumption Research Articles

New System for Simultaneous Measurement of Oxygen Consumption and Changes in Wine Color.

The design, construction and validation of a device for the accurate measurement of the dissolved oxygen content in wine and simultaneously the variation of its spectral fingerprint is presented. The novelty of this system is due to two innovative approaches. First, robustness in measurements is obtained by using cuvettes designed to simultaneously measure the dissolved oxygen and color. Secondly, automatic monitoring is performed to ensure that measurements are always taken at the same cuvette position. The fine-tuning of the device with the study of white and red wines makes it possible, on the one hand, to establish the appropriate measurement conditions and, on the other hand, to determine the amount of oxygen required to cause specific changes in the wine spectrum, information that could not be obtained until now. The preliminary results are very interesting, presenting precise data on the amount of oxygen consumed by the wine and the variations in its visible spectrum, thus reflecting the modification of the responsible phenolic compounds. This information is of great interest, since it helps to optimize the handling of the wine and, if necessary, to moderate the uptake of oxygen in each type of wine to ensure the maintenance of the color during the winemaking and conservation processes of each type of wine. The results of the experiments indicate that this new instrument is feasible and accurate for detecting oxygen changes during wine production.

Proline Oxidation Supports Mitochondrial ATP Production When Complex I Is Inhibited.

The oxidation of proline to pyrroline-5-carboxylate (P5C) leads to the transfer of electrons to ubiquinone in mitochondria that express proline dehydrogenase (ProDH). This electron transfer supports Complexes CIII and CIV, thus generating the protonmotive force. Further catabolism of P5C forms glutamate, which fuels the citric acid cycle that yields the reducing equivalents that sustain oxidative phosphorylation. However, P5C and glutamate catabolism depend on CI activity due to NAD+ requirements. NextGen-O2k (Oroboros Instruments) was used to measure proline oxidation in isolated mitochondria of various mouse tissues. Simultaneous measurements of oxygen consumption, membrane potential, NADH, and the ubiquinone redox state were correlated to ProDH activity and F1FO-ATPase directionality. Proline catabolism generated a sufficiently high membrane potential that was able to maintain the F1FO-ATPase operation in the forward mode. This was observed in CI-inhibited mouse liver and kidney mitochondria that exhibited high levels of proline oxidation and ProDH activity. This action was not observed under anoxia or when either CIII or CIV were inhibited. The duroquinone fueling of CIII and CIV partially reproduced the effects of proline. Excess glutamate, however, could not reproduce the proline effect, suggesting that processes upstream of the glutamate conversion from proline were involved. The ProDH inhibitors tetrahydro-2-furoic acid and, to a lesser extent, S-5-oxo-2-tetrahydrofurancarboxylic acid abolished all proline effects. The data show that ProDH-directed proline catabolism could generate sufficient CIII and CIV proton pumping, thus supporting ATP production by the F1FO-ATPase even under CI inhibition.

A versatile EPR toolbox for the simultaneous measurement of oxygen consumption and superoxide production

In this paper, we describe an assay to analyze simultaneously the oxygen consumption rate (OCR) and superoxide production in a biological system. The analytical set-up uses electron paramagnetic resonance (EPR) spectroscopy with two different isotopically-labelled sensors: 15N-PDT (4-oxo-2,2,6,6-tetramethylpiperidine-d16-15N-1-oxyl) as oxygen-sensing probe and 14N-CMH (1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine, a cyclic hydroxylamine, as sensor of reactive oxygen species (ROS). The superoxide contribution to CMH oxidation is assessed using SOD or PEGSOD as controls. Because the EPR spectra are not superimposable, the variation of EPR linewidth of 15N-PDT (linked to OCR) and the formation of the nitroxide from 14N-CMH (linked to superoxide production) can be recorded simultaneously over time on a single preparation. The EPR toolbox was qualified in biological systems of increasing complexity. First, we used an enzymatic assay based on the hypoxanthine (HX)/xanthine oxidase (XO) which is a well described model of oxygen consumption and superoxide production. Second, we used a cellular model of superoxide production using macrophages exposed to phorbol 12-myristate 13-acetate (PMA) which stimulates the NADPH oxidase (NOX) to consume oxygen and produce superoxide. Finally, we exposed isolated mitochondria to established inhibitors of the electron transport chain (rotenone and metformin) in order to assess their impact on OCR and superoxide production. This EPR toolbox has the potential to screen the effect of intoxicants or drugs targeting the mitochondrial function.

Oxygen uptake during functional activities after stroke-Reliability and validity of a portable ergospirometry system.

BackgroundPeople with stroke have a low peak aerobic capacity and experience increased effort during performance of daily activities. The purpose of this study was to examine test-retest reliability of a portable ergospirometry system in people with stroke during performance of functional activities in a field-test. Secondary aims were to examine the proportion of oxygen consumed during the field-test in relation to the peak-test and to analyse the correlation between the oxygen uptake during the field-test and peak-test in order to support the validity of the field-test.MethodsWith simultaneous measurement of oxygen consumption, participants performed a standardized field-test consisting of five activities; walking over ground, stair walking, stepping over obstacles, walking slalom between cones and from a standing position lifting objects from one height to another. All activities were performed in self-selected speed. Prior to the field-test, a peak aerobic capacity test was performed. The field-test was repeated minimum 2 and maximum 14 days between the tests. ICC2,1 and Bland Altman tests (Limits of Agreement, LoA) were used to analyse test-retest reliability.ResultsIn total 31 participants (39% women, mean (SD) age 54.5 (12.7) years and 21.1 (14.3) months’ post-stroke) were included. The ICC2,1 was ≥ 0.80 for absolute V̇O2, relative V̇O2, minute ventilation, CO2, respiratory exchange ratio, heart rate and Borgs rating of perceived exertion. ICC2,1 for total time to complete the field-test was 0.99. Mean difference in steady state V̇O2 during Test 1 and Test 2 was -0.40 (2.12) The LoAs were -3.75 and 4.51. Participants spent 60.7% of their V̇O2peak performing functional activities. Correlation between field-test and peak-test was 0.689, p = 0.001 for absolute and 0.733, p = 0.001 for relative V̇O2.ConclusionsThis study presents first evidence on reliability of oxygen uptake during performance of functional activities after stroke, showing very good test-retest reliability. The secondary analysis showed that the amount of energy spent during the field-test relative to the peak-test was high and the correlation between the two test was good, supporting the validity of this method.

The cessation of breathing in the chicken embryo during cold-hypometabolism

The avian embryo toward end-incubation combines gas exchange through the chorioallantoic membrane (CAM) and pulmonary ventilation (V˙E). The main experiments examined breathing activity during cold-hypometabolism. Chicken embryos close to hatching were prepared for simultaneous measurements of oxygen consumption (V˙O2) and carbon dioxide production (V˙CO2; open-flow methodology) and breathing frequency (f; barometric technique). As ambient (Ta) and egg temperature (Tegg) dropped, breathing eventually ceased at ∼18°C, when V˙O2 and V˙CO2 were 22–28% of the normothermic values. With the eggshell experimentally covered to reduce CAM gas exchange breathing ceased at slightly lower V˙O2 and V˙CO2 (17–18% of normothermia). Once breathing had stopped, egg exposure to hypoxia (10% or 5% O2) or hypercapnia (3% or 8% CO2) did not resume breathing, which recovered with re-warming. In normothermia, 10% O2 caused hypometabolism and tachypnea; differently, in 5% O2V˙O2 dropped as much as with hypothermia and breathing stopped, to recover upon return in air. Correlation analysis among Ta, Tegg, V˙O2,V˙CO2 and f during cooling and re-warming indicated that f followed more closely the changes in V˙O2 and, especially, in V˙CO2 than the changes in Ta or Tegg. Some considerations suggest that in this experimental model the cessation of breathing in hypothermia or severe hypoxia may be due to hypometabolism, while the lack of chemo-responses may have a different mechanistic basis.

Simultaneous Real-Time Monitoring of Oxygen Consumption and Hydrogen Peroxide Production in Cells Using Our Newly Developed Chip-Type Biosensor Device

All living organisms bear its defense mechanism. Immune cells during invasion by foreign body undergoes phagocytosis during which monocyte and neutrophil produces reactive oxygen species (ROS). The ROS generated in animal cells are known to be involved in several diseases and ailments, when generated in excess. Therefore, if the ROS generated in cells can be measured and analyzed precisely, it can be employed in immune function evaluation and disease detection. The aim of the current study is to introduce our newly developed chip-type biosensor device with high specificity and sensitivity. It comprises of counter electrode and working electrodes I and II. The counter electrode is a platinum plate while the working electrodes I and II are platinum microelectrode and osmium-horseradish peroxidase modified gold electrode, respectively which acts as oxygen and hydrogen peroxide (H2O2) detection sensors. Simultaneous measurement of oxygen consumption and H2O2 generation were measured in animal cells under the effect of exogenous addition of differentiation inducer, phorbol 12-myristate 13-acetate. The results obtained showed considerable changes in reduction currents in the absence and presence of inducer. Our newly developed chip-type biosensor device is claimed to be a useful tool for real-time monitoring of the respiratory activity and precise detection of H2O2 in cells. It can thus be widely applied in biomedical research and in clinical trials being an advancement over other H2O2 detection techniques.

Simultaneous Hypoxia and Low Extracellular pH Suppress Overall Metabolic Rate and Protein Synthesis In Vitro.

BackgroundThe tumor microenvironment is characterized by regions of hypoxia and acidosis which are linked to poor prognosis. This occurs due to an aberrant vasculature as well as high rates of glycolysis and lactate production in tumor cells even in the presence of oxygen (the Warburg effect), which weakens the spatial linkage between hypoxia and acidosis.MethodsFive different human squamous cell carcinoma cell lines (SiHa, FaDuDD, UTSCC5, UTSCC14 and UTSCC15) were treated with hypoxia, acidosis (pH 6.3), or a combination, and gene expression analyzed using microarray. SiHa and FaDuDD were chosen for further characterization of cell energetics and protein synthesis. Total cellular ATP turnover and relative glycolytic dependency was determined by simultaneous measurements of oxygen consumption and lactate synthesis rates and total protein synthesis was determined by autoradiographic quantification of the incorporation of 35S-labelled methionine and cysteine into protein.ResultsMicroarray analysis allowed differentiation between genes induced at low oxygen only at normal extracellular pH (pHe), genes induced at low oxygen at both normal and low pHe, and genes induced at low pHe independent of oxygen concentration. Several genes were found to be upregulated by acidosis independent of oxygenation. Acidosis resulted in a more wide-scale change in gene expression profiles than hypoxia including upregulation of genes involved in the translation process, for example Eukaryotic translation initiation factor 4A, isoform 2 (EIF4A2), and Ribosomal protein L37 (RPL37). Acidosis suppressed overall ATP turnover and protein synthesis by 50%. Protein synthesis, but not total ATP production, was also suppressed under hypoxic conditions. A dramatic decrease in ATP turnover (SiHa) and protein synthesis (both cell lines) was observed when hypoxia and low pHe were combined.ConclusionsWe demonstrate here that the influence of hypoxia and acidosis causes different responses, both in gene expression and in de novo protein synthesis, depending on whether the two factors induced alone or overlapping, and as such it is important for in vivo studies to take this into account.

A novel method for determining human ex vivo submaximal skeletal muscle mitochondrial function.

The present study utilized a novel method aiming to investigate mitochondrial function in human skeletal muscle at submaximal levels and at a predefined membrane potential. The effect of age and training status was investigated using a cross-sectional design. Ageing was found to be related to decreased leak regardless of training status. Increased training status was associated with increased mitochondrial hydrogen peroxide emission. Despite numerous studies, there is no consensus about whether mitochondrial function is altered with increased age. The novelty of the present study is the determination of mitochondrial function at submaximal activity rates, which is more physiologically relevant than the ex vivo functionality protocols used previously. Muscle biopsies were taken from 64 old or young male subjects (aged 60-70 or 20-30 years). Aged subjects were recruited as trained or untrained. Muscle biopsies were used for the isolation of mitochondria and subsequent measurements of DNA repair, anti-oxidant capacity and mitochondrial protein levels (complexes I-V). Mitochondrial function was determined by simultaneous measurement of oxygen consumption, membrane potential and hydrogen peroxide emission using pyruvate + malate (PM) or succinate + rotenone (SR) as substrates. Proton leak was lower in aged subjects when determined at the same membrane potential and was unaffected by training status. State 3 respiration was lower in aged untrained subjects. This effect, however, was alleviated in aged trained subjects. H2 O2 emission with PM was higher in aged subjects, and was exacerbated by training, although it was not changed when using SR. However, with a higher manganese superoxide dismuthase content, the trained aged subjects may actually have lower or similar mitochondrial superoxide emission compared to the untrained subjects. We conclude that ageing and the physical activity level in aged subjects are both related to changes in the intrinsic functionality of the mitochondrion in skeletal muscle. Both of these changes could be important factors in determining the metabolic health of the aged skeletal muscle cell.

Simultaneous Detection of Hydrogen Peroxide Production and Oxygen Consumption By Electrochemical Method in THP-1 Cells during Respiratory Burst

1. Introduction Rapid consumption of oxygen and generation of various reactive oxygen species (ROS), collectively referred to as respiratory burst, occur in immune cells while they are in the processes of bacterial killing and phagocytosis as biological defense mechanisms. It is of significant importance in clinical fields to know the ROS generation mechanism and quantity therein. We have been studying electrochemical means to measure the respiratory burst in THP-1, a cell line from myeloid leukemia patients1) 2). In this study, we aimed to develop an electrochemical sensor for simultaneous and real-time monitoring of the rapid oxygen consumption and hydrogen peroxide generation and to examine the time and amount of hydrogen peroxide generation and oxygen consumption using the sensor. 2 . Experimental 2-1 Composition of 2ch-type electrochemical chip biosensor A chip-type electrochemical sensor was used that comprised working electrode 1 (Pt electrode; f 20 µm) for oxygen detection, working electrode 2 (Os-HRP-modified Au electrode; f 1 mm) for hydrogen peroxide detection, a platinum electrode (1 × 3 mm) as the counter electrode, and a measurement well (internal diameter 5 mm, height 10 mm). 2-2 Measurement conditions THP-1 cell suspension (3.0 × 105 cells/well) was introduced into a well of this sensor, and phosphate buffered saline (PBS; Sigma chemicals, USA) and 11.4 mM glucose solution were used as measurement solutions. Ag/AgCl electrode was used as a control electrode and -0.5 V vs. Ag/AgCl and 0.0 V vs. Ag/AgCl were applied to working electrodes 1 and 2, respectively; the sample was measured for 90 minutes with 1-second sampling time. Twenty minutes after starting the measurement, PMA was added in drops to reach a final concentration of 20 nM or 200 nM, and each reduction current was monitored. 3. Results and Discussion Simultaneous measurements of oxygen consumption and hydrogen peroxide generation in THP-1 using a 2ch-type electrochemical chip biosensor Fig. 1 shows the result of the simultaneous measurements of the oxygen reduction current and hydrogen peroxide reduction current during the respiratory burst in THP-1 cells using a chip-type device. The value of oxygen reduction current dropped immediately after the drop-wise addition of PMA, and 10-20 minutes later, it came back to the same level as before the addition of PMA. This is likely due to the respiratory burst that occurred in THP-1 cells as a result of the rapid consumption of oxygen. Meanwhile, the hydrogen peroxide reduction current increased 15 minutes after starting PMA addition, and continuous generation was observed for approximately 60 minutes. This result suggests that hydrogen peroxide was generated for a long period of time. When it contained no cells, the oxygen reduction current was about -5.5 nA. With this value as a standard (dissolved oxygen concentration 209 µM 3)), the dissolved oxygen concentration during normal respiration and respiratory burst in THP-1 cells was calculated and the generated hydrogen peroxide concentration was further determined from a calibration curve. As a result, the dissolved oxygen concentration during the normal respiration in THP-1 cells was determined to be around 125 µM. On the other hand, the dissolved oxygen concentration during the respiratory burst was determined to be 119 µM and 115 µM when the PMA concentration was 20 nM and 200 nM, respectively. Furthermore, the generated hydrogen peroxide was calculated to be 4.5 nM and 8.3 nM when the PMA concentration was 20 nM and 200 nM. Based on these results, the oxygen consumption and hydrogen peroxide generation during the respiratory burst in THP-1 cells was suggested to be dependent on PMA concentration. Moreover, we confirmed that during the respiratory burst, rapid consumption of oxygen occurs first, and then the generation of hydrogen peroxide starts in a process returning to normal respiration and continues for about one hour. In the future, in addition to the bactericidal effect triggered by PMA stimulation, we plan to examine the respiratory burst and generation of hydrogen peroxide during phagocytosis and conduct more quantitative analysis. References S. Kasai, H. Kikuchi, M. Suzuki, M. Nakano, K. Y. Inoue, T. Honmo, M. Tada, S. Aoyagi, M. Kobayashi and T. Matsue, 225 th ECS meeting (2014). S. Kasai, T. Numata, H. Shiku, H. Abe, T. Matsue, Y. Niizeki, 215 th ECS meeting (2009). H. Shiku, T. Shiraishi, S. Aoyagi, Y. Utsumi, M. Matsudaira, H. Abe, H. Hoshi, S. Kasai, H. Ohya, T. Matsue, Analytica Chimica Acta, 522, (2004), 51–58. Figure 1

Respirometry: Correcting for Diffusion and Validating the Use of Plastic Multiwell Plates with Integrated Optodes

Polystyrene multiwell plates with integrated optodes act as multiple closed-system respirometers that enable the simultaneous measurement of oxygen consumption in small animals. However, the diffusion of oxygen through polystyrene needs to be taken into consideration. Here we provide an equation that accounts for the empirically determined rate of oxygen through a polystyrene well when calculating the instantaneous rate of oxygen consumption. Furthermore, we describe a novel method of calibrating a small respirometer for accuracy using micro-osmotic pumps containing an oxygen scavenger, which is delivered at a constant rate and therefore yields a constant rate of oxygen consumption in an airtight system.

Monitoring breathing in avian embryos and hatchlings by the barometric technique

This communication describes the application of the barometric technique to the measurements of the breathing pattern (tidal volume and breathing rate) and pulmonary ventilation (VE) in chicken embryos and hatchlings. The chamber-plethysmograph was separated into two sections, an animal compartment, maintained at incubation temperature, and a recording compartment kept at a lower temperature. In the case of the embryos, the eggshell itself represented the animal compartment. The hatchlings were accommodated into a "nest" compartment. A flow-through system permitted simultaneous measurements of oxygen consumption (VO2) and carbon dioxide production. Values of breathing rate corresponded to those counted visually in hatchlings while resting in the incubator, and data of resting VE were similar to those obtained by airflow plethysmography, a more invasive technique applicable only to the hatchlings. At rest, the ventilatory equivalent (VE/VO2) of the hatchlings was similar to that reported for adult birds, while in embryos it was much lower. Hypoxia (15 and 10% O2) and hypercapnia (2 and 4% CO2) caused hyperventilation (increase in VE/VO2), both in the hatchlings and in the embryos, these latter using hypometabolism as the main approach to hyperventilate in hypoxia. We conclude that the barometric technique can be adapted to the study of breathing and VE responses in avian embryos and hatchlings.

Discrimination of wheat varieties by simultaneous measurements of oxygen consumption and consistency of flour dough during mixing

AbstractUsing an instrumented mixing reactor, oxygen consumption and torque evolution were recorded during the mixing of wheat flour doughs from eight pure French wheat varieties harvested in 2002 and 2003. Wheat flours were also characterised by biochemical analyses including lipoxygenase (LOX) activity assay, free polyunsaturated fatty acid (PUFA) content, glutathione and ferulic acid contents, results of alveograph and farinograph tests and breadmaking performances. Some parameters defined from the obtained torque curves, equivalent to consistency curves, were correlated with alveograph and farinograph data. The importance of LOX activity levels and free PUFA content to explain the oxygen consumption during mixing is confirmed. In addition to the characteristics of the oxygen consumption curves, the extent to which the consistency curve parameters are able to differentiate among doughs made with flours from different wheat varieties is considered. Copyright © 2006 Society of Chemical Industry

Metabolic and microbial responses in western crossbred and Meishan growing pigs fed a high-fiber diet.

Four Duroc x White composite crossbred (21.8 +/- 1.0 kg BW) and four 12-wk-old Meishan purebred (20.7 +/- 1.6 kg BW) growing barrows were used to determine the relative breed differences in metabolic and microbial responses to a high-fiber diet. The pigs were trained to consume 700 g of a diet containing 35% (as-fed basis) dehydrated alfalfa meal once daily. The pigs' daily intakes of DM, N, GE, NDF, and ADF were 610 g, 16.6 g, 2.64 Mcal, 150 g, and 88 g, respectively. On d 12 after surgical catheterization of the portal vein, ileal vein, and carotid artery, a 3-d total urine and feces collection was conducted. On d 24 after surgery, each pig was placed in an open-circuit calorimeter, and its catheters were connected to a system for simultaneous measurements of oxygen consumption by portal-drained viscera and by whole body, and the net portal absorption of VFA after a 24-h fasting and during a 5-h postprandial period. The VFA measured included acetic, propionic, isobutyric, butyric, isovaleric, and valeric acids. A second 3-d total urine and feces collection was conducted on d 30 after surgery. There were no differences (P = 0.13) between the first and second collections in apparent total-tract digestibility coefficients for nutrients and N retention of pigs. Compared with Duroc x White composite crossbred pigs, Meishan pigs had lower (P = 0.05) apparent digestibility coefficients for DM, N, NDF, hemicellulose, and N retention, but their portal-drained viscera used a greater (P = 0.05) fraction of whole-body oxygen consumption. No differences (P = 0.12) were found between Duroc x White composite crossbred and Meishan pigs in total viable bacteria and cellulolytic bacteria from fecal samples, in vitro digestibility of alfalfa NDF fractions by fecal inocula, whole-body oxygen consumption, net portal absorption of VFA, total energy of absorbed VFA, and the potential of absorbed VFA for meeting the energy needs for whole-body heat production. These results indicate that, in contrast to previous beliefs, the ability of Meishan growing pigs to utilize a high-fiber diet is not superior to that of Duroc x White composite crossbred growing pigs.

Simultaneous measurements of oxygen consumption and ammonia-N excretion in embryos and larvae of marine invertebrates

The quantification of oxygen consumption and ammonia-N excretion rates is essential in determining energy requirements for development of larval invertebrates. In larval energetics, there is a need for accurate and uncomplicated techniques to quantify metabolic rates. A method for simultaneous measurements of oxygen and ammonia-N concentrations is presented. It employs sealed respirometric chambers (ca. 30 ml) in which embryos and larvae are incubated. Analysis is carried out in end-point samples by Winkler's titration and indophenol-blue for oxygen and ammonia-N, respectively. Water is sampled into volume-calibrated glass syringes and oxygen consumption and ammonia-N excretion rates were determined by the difference between experimental and control (no animals) units. The method was successfully used to measure metabolic rates in embryo and larval stages of the shrimp Farfantepenaeus paulensis and in veliger of the mussel Perna perna. The accuracy denoted by the coefficient of variation is comparable to previous results on larval metabolic rates. A biomass: volume (μg ml −1) is proposed to extend its application to further species of marine invertebrates. The method is simple to operate, involves non-expensive material and is portable enough for field work. A substantial number of replicates can be analyzed at the same time and O:N ratio, an indicator of the catabolized substrate, can be calculated.

Postmetamorphic Growth and Metabolism of Long-Spined Black Sea Urchin (Diadema antillarum) Reared in the Laboratory

Postmetamorphic growth and metabolism measurements were obtained on two cohorts of laboratory-reared Diadema antillarum. The cohorts grew linearly from less than 1 mm to over 43 mm. Daily growth averaged 0.097 and 0.11 mm d−1, respectively, for the two cohorts, and was found to differ significantly. Urchin metabolism was examined by a series of simultaneous measurements of oxygen consumption and ammonium excretion over 16 days on starved juveniles ranging 16.5 to 18.3 mm. Metabolic activity under conditions of starvation was used as a test of the viability of urchins reared in the laboratory with cultured food resources. Catabolic activity differed from the first week of starvation compared to the second. Metabolic response included: (1) a 2.2-fold increase in oxygen consumption rate; (2) 50% decline in ammonium excretion rate; and (3) a 5.1-fold increase in oxygen to nitrogen ratio. These measurements are consistent with a shift from almost pure protein catabolism during the first seven days of starvation to a lipid : protein catabolic ratio of 1 : 1 after the first week. Growth and metabolism experiments of this type are seen as a first step towards optimizing laboratory culture techniques of this species.

Hazard identification of pharmaceutical wastewaters using biodegradability studies

A reliable wastewater characterization is an integral part of treatment and management strategies for industrial effluents. This is especially true for the pharmaceutical industry, which exhibits significant differences in its line of activity, generating effluents of very specific and complex natures. Any hazard or risk assessment of wastewater and/or determination of its treatability must include an evaluation of its degradability. Usually various non-standardized laboratory or pilot-scale long-term tests are run by measuring summary parameters for several days to determine the biodegradation potential of the effluent. A complex approach, based on stabilization studies, was proposed to determine the hazardous impact of wastewaters in terms of biodegradable and persistent toxicity. The objective of our work was to carry out complex hazard evaluation of pharmaceutical wastewaters. Whole effluent toxicity was determined using two different toxicity tests. First, we measured the inhibition of oxygen consumption by activated sludge. The test indicated toxicity of the wastewater and thus we performed an additional acute toxicity test with luminescent bacteria Vibrio fisheri. The next step was the determination of whole effluent ready biodegradability. It was determined with simultaneous measurement of oxygen consumption (ISO 9804) and carbon dioxide production (ISO 9439) in a closed respirometer, accompanied by DOC/IC measurements. The pharmaceutical wastewater degraded readily (83%, lag phase was 2 days, biodegradation rate was 0.33999 day(-1)) on the basis of O2 measurements. The biodegradation, calculated from the CO2 measurements, was comparable. We also applied mass balances of DOC/IC at the beginning and at the end of biodegradation experiments to confirm the extent and rate of biodegradation. The determination of hazardous impact and treatability of the effluent was concluded with aerobic stabilization studies. Biodegradation of the wastewater during the study was followed by relevant biochemical analysis and DOC/IC mass balance.

Effects of training on respiratory gas exchange, nitrogenous waste excretion, and fuel usage during aerobic swimming in juvenile rainbow trout (Oncorhynchus mykiss)

Fuel utilization during aerobic exercise was determined in juvenile rainbow trout that had previously undergone 2 weeks of continuous aerobic training at 25% of their maximum sustainable speed (Ucrit) and was compared with that in untrained trout. Instantaneous fuel usage was calculated from simultaneous measurements of oxygen consumption ( MO2), carbon dioxide excretion ( MCO2), and nitrogenous waste excretion (MN). Over 58 h of sustained aerobic exercise at 55% of Ucrit, MO2and MCO2remained virtually constant in the trained fish, in contrast to the significant fall over time in untrained fish, but total gas exchange was similar in the two groups. Aerobic respiratory quotient remained constant in the two treatments. MNwas lower in the trained fish, resulting in a much lower (and stable) nitrogen quotient; the nitrogen quotient increased with swimming duration in the untrained fish. Lipid was the major fuel source powering aerobic exercise in both groups. Carbohydrate also played a significant role, whereas protein was of lowest quantitative importance. Training resulted in a decrease in reliance on protein and an increase in reliance on lipid metabolism. The role of protein oxidation in supporting aerobic swimming in fish, which is smaller than commonly believed, is further reduced by training.

Effects Of Ca2+ On Oxidative Phosphorylation In Mitochondria From The Thermogenic Organ Of Marlin

Mitochondria from the muscle-derived thermogenic (heater) organ and oxidative red muscle of the blue marlin (Makaira nigricans) were studied in order to evaluate aspects of the mechanism of thermogenesis in heater tissue. We investigated whether short-term Ca(2+)-induced uncoupling of mitochondria contributes to the thermogenic cycle of the heater organ by enhancing the respiration rate. Specific electrodes were used to obtain simultaneous measurements of oxygen consumption and Ca2+ fluxes on isolated mitochondria, and the effects of various concentrations of Ca2+ on respiration rates and the ADP phosphorylated/atomic oxygen consumed (P/O) ratio were examined. Addition of Ca2+ in excess of 10 mumol l-1 to respiring heater organ or red muscle mitochondria partially inhibited state 3 respiration and reduced the P/O ratio, indicating that the mitochondria were partially uncoupled. These effects were blocked by 2 mumol l-1 Ruthenium Red. In heater organ mitochondria, state 3 respiration rate and the P/O ratio were not significantly reduced by 1 mumol l-1 free Ca2+, a concentration likely to be near the maximum achieved in a stimulated cell. This indicates that transient increases in cytosolic Ca2+ concentration may not significantly reduce the P/O ratio of heater organ mitochondria. The activity of 2-oxoglutarate dehydrogenase in heater organ mitochondria was stimulated by approximately 15% by Ca2+ concentrations between 0.2 and 1 mumol l-1. These results suggest that heater organ mitochondria are able to maintain a normal P/O ratio and should maintain ATP output during transient increases in Ca2+ concentration, supporting a model in which an ATP-consuming process drives thermogenesis. Activation of mitochondrial dehydrogenases by low levels of Ca2+ may also enhance respiration and contribute to thermogenesis.

Influence of Joint Impairments on Walking in Children with Cerebral Palsy.

The restriction of the range of motion (ROM) of joints in patients with cerebral palsy (CP) is caused by joint contracture and muscle shortening due to the effects of their disease on the brain. The aim of this study is as to identify the influence of the restricted ROM of joints on walking ability in CP children by means of simultaneous measurements of oxygen consumption, acceleration of body in the vertical direction, stride and cadence during walking on a treadmill. The stride and cadence in CP children were similar to those in normal children. The CP children demonstrated greater oxygen consumption and acceleration compared with normal subjects ; in particular, this trend was more marked in CP group members with restriction of joint movement. These findings suggest the joint impairment contributes to walking disability as well as dysfunction in the neuromuscular system. When the motion of the ankle joints in normal children was restricted with a pair of special short braces, the children exhibited the kind of impaired walking seen in CP children, although the degree of the disability was not as great as that in CP patients. These results also seem to support the close relationship between the joint impairment and walking disability.

Some like it hot calorimetric investigations of voodoo lilies

An isoperibolic calorimeter of approximately 3 liters volume and a sensitivity of 52.8 mV W is described, specially designed for studies on whole plants. It was used to monitor thermogenesis during anthesis (flowering up) of the voodoo lily, Sauromatum guttatum, a member of the family of Araceae. Simultaneous measurements of oxygen consumption indicate a high ratio of heat production to oxygen consumption and a strong uncoupling of oxydative phosphorylation during anthesis. Thermographic registration and temperature determinations along the whole plant show an elevation of the appendix temperature of 7°C above the surrounding.