Initial Rate Of Oxygen Consumption Research Articles

Measuring and Modeling Oxygen Transport and Consumption in 3D Hydrogels Containing Chondrocytes and Stem Cells of Different Tissue Origins.

Understanding how the local cellular environment influences cell metabolism, phenotype and matrix synthesis is crucial to engineering functional tissue grafts of a clinically relevant scale. The objective of this study was to investigate how the local oxygen environment within engineered cartilaginous tissues is influenced by factors such as cell source, environmental oxygen tension and the cell seeding density. Furthermore, the subsequent impact of such factors on both the cellular oxygen consumption rate and cartilage matrix synthesis were also examined. Bone marrow derived stem cells (BMSCs), infrapatellar fat pad derived stem cells (FPSCs) and chondrocytes (CCs) were seeded into agarose hydrogels and stimulated with transforming growth factor-β3 (TGF- β3). The local oxygen concentration was measured within the center of the constructs, and numerical modeling was employed to predict oxygen gradients and the average oxygen consumption rate within the engineered tissues. The cellular oxygen consumption rate of hydrogel encapsulated CCs remained relatively unchanged with time in culture. In contrast, stem cells were found to possess a relatively high initial oxygen consumption rate, but adopted a less oxidative, more chondrocyte-like oxygen consumption profile following chondrogenic differentiation, resulting in net increases in engineered tissue oxygenation. Furthermore, a greater reduction in oxygen uptake was observed when the oxygen concentration of the external cell culture environment was reduced. In general, cartilage matrix deposition was found to be maximal in regions of low oxygen, but collagen synthesis was inhibited in very low (less than 2%) oxygen regions. These findings suggest that promoting an oxygen consumption profile similar to that of chondrocytes might be considered a key determinant to the success of stem cell-based cartilage tissue engineering strategies.

Antioxidant efficiency and mechanisms of green tea, rosemary or maté extracts in porcine Longissimus dorsi subjected to iron-induced oxidative stress

Plant extracts from rosemary (RE), green tea (GTE), and maté (ME) were compared for the protection against iron-induced oxidation in porcine homogenates at total phenolic concentrations from 25 to 250 ppm. Lipid oxidation as indicated by TBARS was in all cases sufficiently suppressed, especially for RE. Hydrophobic RE retarded overall oxidation in the homogenates with an inverted dose-dependent response. Optimum delay of oxygen consumption was found at the lowest concentration applied, similar to protection against thiols and formation of protein radicals as measured by ESR, whereas the high concentration increased oxygen consumption and caused additionally thiol loss possibly due to thiol-quinone interactions, generating protein-phenol complexes. Hydrophilic ME or GTE increased the initial oxygen consumption rate as an indication of prooxidant activities at elevated concentrations. However, they were found to protect myoglobin and protein at those high concentrations with GTE being more efficient, possibly due to better chelation effect.

Evaluation of bioremediation of oil-polluted soil using the respirometric oxitop® method.

Bioremediation includes natural processes to degrade harmful compounds in theenvironment. The goal in bioremediation is to stimulate microorganisms with differentadditives that will activate them to destroy the contaminants. Respirometry methods havebeen used to assess the microbial activity of soil but a few respirational studies have beenperformed with surfactants.The objective of this study was to examine the effects of surfactants and adsorbents onoil-contaminanted soil respirational activity. The measurement of oxygen consumption isused to evaluate the enhancement of the bioremediation of polluted soil by surfactantsand adsorbents. Selected surfactants were sodium dodecyl sulfate (SDS), Saponine andemulsifying agent SR-100. In the present study the respirometric OxiTop® system wasused to measure the oxygen consumption of the soil by adding the different surfactants.The rates of oxygen consumption were calculated from respiration data.The measurements of respiration showed that the maximum initial rate of oxygenconsumption of soil was 4.39 mg 02 kg -1 h -1 at the water content of9% (w/w ). SR-100addition showed the highest initial rate of oxygen consumption - 18.61 mg 02 kg ·1 h -1•The medium rate of oxygen consumption of the oil contaminated soil for five days washalf of the initial rate. The mixtures of soil and the adsorbents had lower respirationalactivity, even when solutions of the surfactants were added. The highest initial rates ofoxygen consumption of soil were 4.24 mg Oi kg -1 h -1 ( Saponine solution was added) and13.00 mg 02 kg"1 h"1 (SR-100 solution was added) for the hydrophobic and hydrophilicadrorbents, respectively. For the mixture of soil and adsorbent the longer lag-period forthe activation of oxygen consumption occurred by the adding of surfactants.

Comparison of PDT parameters for RIF and H460 tumor models during HPPH-mediated PDT.

Singlet oxygen (1O2) is the major cytotoxic species producing PDT effects, but it is difficult to monitor in vivo due to its short life time in real biological environments. Mathematical models are then useful to calculate 1O2 concentrations for PDT dosimetry. Our previously introduced macroscopic model has four PDT parameters: ξ, σ, β and g describing initial oxygen consumption rate, ratio of photobleaching to reaction between 1O2 and cellular targets, ratio of triplet state (T) phosphorescence to reaction between T and oxygen (3O2), and oxygen supply rate to tissue, respectively. In addition, the model calculates a fifth parameter, threshold 1O2 dose ([1O2]rx,sd). These PDT parameters have been investigated for HPPH using radiation-induced fibrosarcoma (RIF) tumors in an in-vivo C3H mouse model. In recent studies, we additionally investigated these parameters in human non-small cell lung carcinoma (H460) tumor xenografts, also using HPPH-mediated PDT. In-vivo studies are performed with nude female mice with H460 tumors grown intradermally on their right shoulders. HPPH (0.25 mg/kg) is injected i.v. at 24 hours prior to light delivery. Initial in vivo HPPH concentration is quantified via interstitial HPPH fluorescence measurements after correction for tissue optical properties. Light is delivered by a linear source at various light doses (12-50 J/cm) with powers ranging from 12 to 150 mW per cm length. The necrosis radius is quantified using ScanScope after tumor sectioning and hematoxylin and eosin (H&E) staining. The macroscopic optimization model is used to fit the results and generate four PDT parameters. Initial results of the parameters for H460 tumors will be reported and compared with those for the RIF tumor.

Comparative oxygen consumption rates of subitaneous and delayed hatching eggs of the calanoid copepod Acartia tonsa (Dana)

Life history strategies can be adaptations to existence in e.g. unstable environments. Under stressful environmental conditions mobile species tend to migrate to more suitable places, but migration in time by dormancy is another durable strategy. More than 50 species of calanoid copepods can produce diapause eggs requiring a long refractory phase. Delayed hatching eggs (DHE; maternally determined oligopause eggs hatching within >72h to a month) have been described as a state between diapause and subitaneous (hatching within 24–72h) because of their very short refractory phase. We used nanorespirometry to monitor initial oxygen consumption rate of individual eggs of the ubiquitous neritic calanoid copepod Acartia tonsa to distinguish between subitaneous and DHE. We hypothesized that subitaneous eggs exhibit higher initial oxygen consumption rates than DHE, and that initial egg oxygen consumption rate is correlated to the time for the individual egg to hatch. Subitaneous eggs exhibited higher initial oxygen consumption rates than DHE and there were no pattern in initial oxygen consumption rates vs. time to hatch or die from the eggs. Variability in initial oxygen consumption rates within batches of both subitaneous and DHE, as well as between these egg types, is prevalent. There was a continuum from sluggish- to fast metabolising eggs considering initial oxygen consumption rates most likely reflecting phenotypic variation within cohorts. No matter the individual initial egg oxygen consumption rate, embryogenesis took place with unpredicted rates and most eggs eventually hatched. Based on our observations and theoretical considerations we suggest that DHE follow a U shape metabolism pattern with time as described for true diapause eggs. DHE strategy (oligopause) is an important life history trait allowing the spread of hatching over a period of time (a month or two), increasing the likelihood that some offspring encounter suitable environmental conditions and can contribute to the pelagic population of neritic calanoids in environmental fluctuating environments.

Reductive activation of terpenylnaphthoquinones

Four terpenylnaphthoquinones were found to enhance the rate of superoxide production in the presence of ascorbate as detected from the superoxide dismutase (SOD)-inhibitable initial oxygen consumption rates. Initial rates of oxygen consumption in the presence of ascorbate plus quinone increase with an increase in the half-wave reduction potentials of the quinones. These quinones also enhance the rate of Cyt(III) c reduction by xanthine/xanthine oxidase (X/XO) in both air- and nitrogen-saturated aqueous solutions at pH 7.4. Maximum rates of Cyt(III) c reduction in nitrogen and oxygen-saturated solutions ( V max), in the presence of X/XO, increase with an increase in the half-wave reduction potentials of the quinones. SOD inhibits Cyt(III) c reduction rates in the presence of these quinones and X/XO in a manner which is also dependent on the quinone half-wave redox potential. The relative antineoplastic activity of two of these quinones follows the order in rates of oxygen consumption or Cyt(III) c reduction. This is consistent with an antineoplastic action of these quinones through the mechanism of redox cycling or possible interference or inhibition of mitochondrial respiration.

KINETICS OF ASPEN WOOD OXIDATION BY OXYGEN IN ALKALINE MEDIA

The kinetics of aspen wood oxidation has been studied using the method of initial rates of oxygen consumption. Under certain conditions, the mass transfer processes practically did not limit the process rate. The reaction orders with respect to oxygen and hydroxide anion were determined experimentally. They depend on the conditions of the reaction, which makes it possible to distinguish the regions of different radical oxidation mechanisms: short chain, long chain and degenerate branching chain.

I: Negative effect of cold ischemia on initial renal function.

Correlation between post-transplant function and exposure to cold ischemia (CI) during preservation has been reported. We attempted to identify the effect of CI on renal function using exsanguinous metabolic support (EMS) technology, to eliminate effects of reperfusion complications. Small bovine kidneys were used to evaluate 4 vs. 24 hours of CI, after warm ischemic (WI) exposure of <15, 30 or 60 minutes. After CI, kidneys were warm perfused (30 degrees C to 32 degrees C) ex vivo using EMS technology. Restored renal metabolism and function were quantified by oxygen consumption, urine production, glomerular filtration rate (GFR), and hemodynamic characteristics. The results demonstrate a CI-associated lag phase in the restoration of metabolism, in which the longer cold-preserved kidneys exhibit a lower initial rate of oxygen consumption. However, after 3 hours of EMS perfusion there was no significant difference in the O2 consumed, urine flow, GFR, perfusion flow, or pressure between the kidneys stored for 4 or 24 hours. An initial reduction in metabolism after longer CI may influence the severity of actual reperfusion injury during transplantation. Therefore, these results provide preliminary evidence suggesting that an acellular warm temperature reperfusion ex vivo may enhance restoration of cellular metabolism and minimize damage from the cold seen upon actual reperfusion.

Kinetics of redox interaction between substituted quinones and ascorbate under aerobic conditions

One-electron reduction of quinones (Q) by ascorbate (AscH −); (1) AscH −+Q→Q − +Asc − +H +, followed by the oxidation of semiquinone (Q − ) by molecular oxygen; (2) Q − +O 2→Q+O 2 − , results in the catalytic oxidation of ascorbate (with Q as a catalyst) and formation of active forms of oxygen. Along with enzymatic redox cycling of Q, this process may be related to Q cytotoxicity and underlie an antitumor activity of some Qs. In this work, the kinetics of oxygen consumption accompanied the interaction of ascorbate with 55 Qs including substituted 1,4- and 1,2-benzoquinones, naphthoquinones and other quinoid compounds were studied in 50 mM sodium phosphate buffer, pH 7.40, at 37°C by using the Clark electrode technique. The capability of Q to catalyze ascorbate oxidation was characterized by the effective value of k EFF calculated from the initial rate of oxygen consumption ( R OX) by the equation R OX= k EFF[Q][AscH −] as well as by a temporary change in R OX. The correlation of k EFF with one-electron reduction potential, E(Q/Q − ), showed a sigma-like plot, the same for different kinds of Qs. Only the Qs which reduction potential E(Q/Q − ) ranged from nearly −250 to +50 mV displayed a pronounced catalytic activity, k EFF increased with shifting E(Q/Q − ) to positive values. The following linear correlation between k EFF (in M −1 s −1) and E(Q/Q − ) (in mV) might be suggested for these Qs: lg( k EFF)=3.91+0.0143 E(Q/Q − ). In contrast, Qs with E(Q/Q − )<−250 mV and E(Q/Q − )>+50 mV showed no measurable catalytic activity. The Qs studied displayed a wide variety in the kinetic regularities of oxygen consumption. When E(Q/Q − ) was more negative than −100 mV, Q displayed a simple (‘standard’) kinetic behavior— R OX was proportional to [AscH −][Q] independently of concentration of individual reagents, [AscH −] and [Q]; R OX did not decrease with time if [AscH −] was held constant; Q recycling was almost reversible. Meanwhile, Qs with E(Q/Q − )>−100 mV demonstrated a dramatic deviation from the ‘standard’ behavior that was manifested by the fast decrease in R OX with time and non-linear dependence of even starting values of R OX on [Q] and [AscH −]. These deviations were caused basically by the participation of Q − in side reactions different from (2). The above findings were confirmed by kinetic computer simulations. Some biological implications of Q–AscH − interaction were discussed.

NADPH and ferredoxin:NADP+ oxidoreductase-dependent reduction of quinones and their reoxidation11Preliminary results were presented at the 10th FESPP Congress, Florence, Italy (Plant Physiol. Biochem., Special issue, 1996, p. 99).

Abstract Molecular oxygen uptake was initiated by adding NADPH (1 mM) to the buffered medium containing 0.6 μ M spinach ferredoxin:NADP + oxidoreductase and 20 μ M quinone (plastoquinone-2, decyl-plastoquinone, decyl-ubiquinone, or duroquinone). At pH 7.7 the rate of oxygen uptake was 2- to 12-fold higher during an initial phase (V 1 ) than in a subsequent phase (V 2 ). Except for duroquinone, the initial rate of oxygen consumption was ca. 2.7-fold higher in alkaline than in acidic medium. Ferredoxin was not essential, although it stimulated the reaction investigated. Oxygen uptake was not detectable with plastoquinone-9 or α-tocoquinone. The possible mechanisms of the NADPH and ferredoxin:NADP oxidoreductase dependent reduction of some quinones and their reoxidation are discussed.

Kinetics of redox interaction between substituted 1,4-benzoquinones and ascorbate under aerobic conditions: Critical phenomena

Redox cycling is believed to be the most general molecular mechanism of quinone (Q) cytotoxicity. Along with redox cycling induced by a reductase, a similar process is known to occur via electron transfer from ascorbate (AscH-) to Q with formation of a semiquinone radical (Q·-):(1) Q+AscH—→Q·—+AsH·—+H+ (2)Q·—+ Q2→Q+2The net effect of reactions (1) and (2) provides for the catalytic oxidation of AscH-, with Q serving as a catalyst. In this work, the kinetics of oxygen consumption accompanying this process were studied with several substituted 1,4-benzoquinones (BQ) at 37°C in phosphate buffer, pH 7.40, using the Clark electrode technique. The value of k1 determined from the initial rate of oxygen consumption was typically found to increase when the one-electron reduction potential E(Q/Q·-) shifted to more positive values. With Q for which E(Q/Q·-) is less than -100 mV, the rate of oxygen uptake (ROX) was found to be directly correlated with the [Q][AscH-] value independent of the concentration of individual reagents, remaining constant for a long period. With mono- and dialkyl-substituted 1,4-BQs, for which E(Q/Q·-) is higher than -100mV, significant deviations from the above simple kinetic regularities were observed. In particular, ROX decreased dramatically with time and critical phenomena (the existence of certain concentrations of Q and/or AscH- above or below which the catalytic oxidation of AscH- ceased completely after a non-stationary period of short duration) were observed. These abnormalities can be explained on the basis of the kinetic scheme which contains, in addition to reactions (1)and (2), several side reactions including that between Q·- and AscH-. Implications of critical phenomena discovered in this study for the problems of Q toxicity and vitamin C avitaminosis are discussed.

Kinetics of Nitrosomonas europaea at extreme substrate, product and salt concentrations

Nitrification of ammonia in concentrated waste streams is gaining a lot of attention nowadays. Nitrosomonas europaea is the predominant ammonia-oxidizing species in these environments. Prediction of the behaviour of a pure culture of N. europaea (ATCC 19718) under conditions prevailing in concentrated waste streams was the aim of this study. The initial oxygen consumption rate of a concentrated cell suspension was used as a rapid assay to measure the effects on N. europaea under various conditions. Several relationships, based on Michaelis-Menten kinetics, were derived. They describe the behaviour of N. europaea at substrate (NH4 +), product (NO2 − and K+, Na+, SO inf4 sup2− , NO3 −, Cl−) concentrations up to 500 mol/m3 and pHs ranging from 6.5 to 8.5. High concentrations of ions inhibited N. europaea but specific substrate inhibition was not observed. Product inhibition was strongly pH-dependent and severe inhibition was found at pH 6.5.

Maintenance of motility of fowl spermatozoa in vitro is prolonged by a low molecular weight factor derived from cultured chick embryo cells.

Gel filtration of a conditioned medium composed of the supernatant fluid removed from a 5-day culture of skeletal muscle cells from 9-day-old chick embryos with Bio-Gel P-2 revealed one peak of motility-prolonging activity (about 0.3 kDa), which was not present in fresh medium. Spermatozoa incubated in this fraction of the conditioned medium maintained their motility for at least 36 h at 37 degrees C. Both the formation of lipid peroxide and the leakage of lactic dehydrogenase of spermatozoa incubated in the conditioned medium fraction were lower than those incubated in the corresponding fresh medium. Initial rate of oxygen consumption of the spermatozoa incubated in the conditioned medium fraction increased compared with that of the fresh medium fraction. These results suggest that a low molecular weight factor(s) supplied by cultured cells effectively prolongs the motility of fowl spermatozoa, and that the effect could result from inhibition of the structural damage to the sperm membrane.

Reducing the influence of sample oxygen in biosensor design

This paper deals with principles of designing enzyme biosensors in which the effect of sample oxygen is reduced. A glucose sensor was developed consisting of two chambers for the sample and electrolyte solution, a semi-permeable membrane dividing them, enzyme (glucose oxidase) dissolved in the electrolyte solution and an oxygen electrode. The behaviour of the biosensor was studied by analysis of a three-layer dynamic model and experimental investigations. It is demonstrated that the influence of sample oxygen is reduced when oxygen needed for the enzymatic reaction is supplied from the electrolyte solution and when transport of glucose and oxygen through the membrane is restricted. An additional decrease in this effect is also provided by choosing an optimal chamber volume ratio and by utilizing initial rates of oxygen consumption for detecting substrate concentration. The increase in the first derivative of the glucose sensor's current at pO 2 = 40 mm Hg was about 2–3% compared with its value at pO 2 = 145 mm Hg for samples containing 5 mM glucose. The operation of the biosensor under these conditions also provided greater measurement precision, an increase in the linear range of the calibrated curve and independence of measurements from a decrease in enzymatic activity. The calibrated curve of the glucose sensor was linear in the range 0·1–25 mM. The accuracy of measurements within this range was ± 1·8% and the relative standard deviation was ± 1·3% at 5 mM. The manufactured glucose sensor is noted for many advantages, including rapid measurements and that no operation for the preparation of a sample or its dosage is necessary. The measurement time did not exceed 30 s. Only 20 μl of whole blood and 100 μl of enzyme solution are required for a single analysis. The biosensor remained stable for more than eight months at a storage temperature for the enzyme solution below 20–25°C.

Individual variations in the response to hypoxia inCancer pagurus(L.) measured at the excited rate

The excited rates of oxygen consumption in individual Cancer pagurus were measured at 78% and 54% levels of O2 saturation, at ambient temperature (16°C) and normal salinity. From these measurements, the ability to compensate for mild hypoxia was calculated. The wet wts of the entire crab, and of the digestive gland, the dry wts of the gills, and the wet wt/dry wt ratio of one of the claws were recorded. The relative size‐free values of oxygen consumption, gill wt, digestive gland wt, and the dry wt/wet wt ratio were all positively correlated with one another, thus the crabs are metabolically and morphologically symmetrical; as well as individually different. There was on average, no ability to compensate for reduced oxygen tension (‐6%) when the crabs were excited, but the significant ability to compensate in individual crabs was inversely proportional to their initial rate of oxygen consumption, as was found earlier in Carcinus maenas. There was a very highly significant individuality in these crabs, expressed as correlations between their organ sizes and physiological abilities.

Site-specific DNA damage induced by sulfite in the presence of cobalt(II) ion: Role of sulfate radical

The reactivities of sulfite (SO 3 2−) with DNA in the presence of metal ions were investigated by a DNA sequencing technique using 32P-labeled DNA fragments obtained from human c-Ha- ras-1 protooncogene. Sulfite caused DNA damage in the presence of Co 2+, Cu 2+ and Mn 2+, although sulfite alone or metal ion alone did not. The order of inducing effect on sulfite-dependent DNA damage (Co 2+ > Cu 2+ > Mn 2+ > Fe 3+) was consistent with that of accelerating effect on the initial oxygen consumption rate of sulfite autoxidation. The DNA damage induced by sulfite plus Co 2+ was inhibited by 3,5-dibromo-4-nitrobenzenesulfonate, primary and secondary alchols, whereas it was not inhibited by SOD, catalase and tert-butyl alcohol. Incubation of DNA with sulfite plus Co 2+ followed by the piperidine treatment led to the predominant cleavage at the positions of guanine especially located 5' to guanine. Sulfite plus Cu 2+ gave a DNA cleavage pattern different from that induced by sulfite plus Co 2+. The photolysis of peroxydisulfate (S 8O 2 2−), which is known to produce SO 4 − radicals, gave a DNA cleavage pattern similar to that induced by sulfite plus Co 2+. ESR studies using spin-trapping reagent revealed the production of spin adduct possibly of SO 3 − radical in a solution of sulfite plus Cu 2+, whereas much less spin adduct was produced by sulfite plus Co 2+. The results suggest that sulfite is rapidly autoxidized in the presence of Co 2+ to produce SO 4 − radical causing site-specific DNA damage.

Epoxidation of 7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene via a hydroperoxide-dependent mechanism catalyzed by lipoxygenases

The lipoxygenase catalyzed epoxidation of 7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (BP-7,8-diol) was examined. Epoxidation of the BP-7,8-diol was catalyzed by 5- and 15-lipoxygenase in the presence of either arachidonic acid, gamma-linolenic acid, or 15-hydroperoxyeicosatetraenoic acid (15-HPETE). The anti-9,10-epoxy-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene isomer was formed in greater quantities than the syn isomer, indicative of peroxyl radical mediated epoxidation. Epoxidation was dependent on time, enzyme and fatty acid concentration. There was no difference in the time course of epoxidation with either arachidonic acid or 15-HPETE, although the initial rate of oxygen consumption was approximately 55-fold greater with arachidonic acid. The lipoxygenase inhibitor and anti-oxidant nordihydroguaiaretic acid inhibited epoxidation in a dose-dependent manner in incubations initiated with either arachidonic acid or 15-HPETE. The anti-oxidant butylated hydroxyanisole also inhibited the epoxidation. Incubations conducted under anaerobic conditions with 15-lipoxygenase and either arachidonic acid or 15-HPETE significantly decreased epoxidation. This suggests that the oxygen inserted into BP-7,8-diol is derived from the atmosphere. The epoxidizing peroxyl radicals could not be detected but their precursors, carbon-centered radicals, were detected by using the ESR spin trapping technique in incubations of 15-lipoxygenase with 15-HPETE. This radical, formed by reduction and rearrangement of the hydroperoxide, may trap oxygen to form a peroxyl radical. We propose that the epoxidizing species is a peroxyl radical derived from 15-HPETE rather than from arachidonic acid. This proposal is based on the similar amounts of epoxidation, but dissimilar amount of oxygen consumed with both fatty acids. Since lipoxygenases are widely distributed in vivo, especially in areas where tumors arise such as the pulmonary epithelium, peroxyl radical formation by these enzymes may have an important role in chemical carcinogenesis.

Effects of alpha-tocopherol, its carboxylic acid chromane compound and two novel antioxidant isoflavanones on prostaglandin H synthase activity and autodeactivation.

The natural antioxidant alpha-tocopherol has repeatedly been described to inhibit platelet aggregation and thromboxane formation, whereas its influence on prostaglandin H synthase in vivo and in vitro is a matter of controversy. In the present study the effects of different antioxidative compounds on ram vesicular gland microsomal prostaglandin H synthase activity were investigated in vitro: d,l-alpha-tocopherol, its carboxylic acid chromane compound (Trolox), phytol, alpha-tocopherol-acetate and two novel antioxidative isoflavanones, obtained by methylation and/or hydrogenation of naturally occurring isoflavones from fermented soybeans (6,7-dihydroxy-4'-methoxyisoflavanone and 6,7,4'-trihydroxyisoflavanone). Alpha-tocopherol, -acetate and phytol revealed no significant influence on the enzyme activity when applied in concentrations up to 1 mM. Trolox (100-1000 mu mumol/l) and the two isoflavanones (5-50 and 10-100 mumol/l) dose-dependently augmented the initial rate of oxygen consumption and the total oxygen uptake during prostaglandin H synthase incubation with arachidonic acid (AA). In parallel, these compounds increased the formation of prostaglandin E2 and F2 alpha from 14C-labelled AA, and they markedly protected the prostaglandin H synthase from rapid autodeactivation as revealed by repetitive application of AA in small doses. We suggest that these compounds serve as cosubstrates to which the oxidizing equivalents are transferred which arise during the hydroperoxidase reaction of the enzyme.

Redox cycling of anthracyclines by cardiac mitochondria. II. Formation of superoxide anion, hydrogen peroxide, and hydroxyl radical.

In the accompanying paper (Davies, K. J. A., and Doroshow, J. A. (1986) J. Biol. Chem. 261, 3060-3067), we have demonstrated that anthracycline antibiotics are reduced to the semiquinone form at Complex I of the mitochondrial electron transport chain. In the experiments presented in this study we examined the effects of doxorubicin (Adriamycin), daunorubicin, and related quinonoid anticancer agents on superoxide, hydrogen peroxide, and hydroxyl radical production by preparations of beef heart submitochondrial particles. Superoxide anion formation was stimulated from (mean +/- S.E.) 1.6 +/- 0.2 to 69.6 +/- 2.7 or 32.1 +/- 1.5 nmol X min-1 X mg-1 by the addition of 90 microM doxorubicin or daunorubicin, respectively. However, the anthracycline 5-iminodaunorubicin, in which an imine group has been substituted in the C ring quinone moiety, did not increase superoxide production over control levels. In the presence of rotenone, initial rates of oxygen consumption and superoxide formation were identical under comparable experimental conditions. Furthermore, H2O2 production increased from undetectable control levels to 2.2 +/- 0.3 nmol X min-1 X mg-1 after treatment of submitochondrial particles with doxorubicin (200 microM). The hydroxyl radical, or a related chemical oxidant, was also detected after the addition of an anthracycline to this system by both ESR spectroscopy using the spin trap 5,5-dimethylpyrroline-N-oxide and by gas chromatographic quantitation of CH4 produced from dimethyl sulfoxide. Hydroxyl radical production, which was iron-dependent in this system, occurred in a nonlinear fashion with an initial lag phase due to a requirement for H2O2 accumulation. We also found that two quinonoid anti-cancer agents which produce less cardiotoxicity than the anthracyclines, mitomycin C, and mitoxantrone, stimulated significantly less or no hydroxyl radical production by submitochondrial particles. These experiments suggest that injury to cardiac mitochondria which is produced by anthracycline antibiotics may result from the generation of the hydroxyl radical during anthracycline metabolism by NADH dehydrogenase.

Limitations of evaluation methods for inhibited oxidation processes in the liquid phase

The limitations of the different evaluation methods used for liquid-phase oxidation processes in the presence of inhibitors have been investigated using computer simulation techniques. The reaction InH + HROOH has been included in a generally accepted mechanism. According to the calculations the rate of initiation and rate coefficient of the reaction InH + HROOH should be determined based on inhibition-period data rather than on values of initial rates of oxygen consumption. Using the same data set different formulae have been suggested depending on the parameters to be calculated.