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Biomedical subjects

E Cadenas

Publications and source records attributed to E Cadenas.

At least 109 records · Page 6Linked to original sources
PubMed1985

Oxidative stress: excited oxygen species and enzyme activity.

The metabolic role of aldehydes, hydroperoxides, and quinones was investigated with emphasis on oxidative transitions involving oxygen free radicals and associated with enzymatic activities. The oxidative metabolism of aldehydes (originating either from ethanol oxidation, or monoamine oxidase activity, or oxidative breakdown of lipid hydroperoxides during lipid peroxidation) is a source of alkane production and low-level chemiluminescence. Since both parameters reflect cellular oxidative conditions, it can be inferred that side-products of aldehyde oxidase activity might participate in the link between the initial enzymatic oxidation of aldehyde and the occurrence of oxidizing species leading to chemiluminescence and alkane production. The metabolism of hydroperoxides was considered under two different aspects: first, the hydroperoxide reduction, within the frame of a detoxication mechanism, as mediated by a selenoorganic compound PZ-51 that displays glutathione peroxidase-like activity and an antioxidant activity; second, the enzyme-catalyzed disproportionation of hydroperoxides as a source of a potent oxidizing equivalent, singlet molecular oxygen. The cytotoxicity of quinones, utilized in therapeutic agents such as anticancer drugs, is believed to be related to oxidative stress due to the formation of the superoxide radical and subsequent more reactive oxygen species. The enzyme-catalyzed one-electron reduction of menadione seems to play a substantial role in the development of cytotoxic effects, at variance with the 2-electron reduction of the quinone. The observation of low-level chemiluminescence under conditions which favor the one-electron reduction process or which diminished the two-electron reduction process indicates the practicability of low-level chemiluminescence measurements in monitoring changes in quinone metabolism and related cytotoxic effects.

Aldehydes↗
PubMed1984

Evaluation of alpha-tocopherol antioxidant activity in microsomal lipid peroxidation as detected by low-level chemiluminescence.

The significance of microsomal vitamin E in protecting against the free-radical process of lipid peroxidation was evaluated with the low-level-chemiluminescence technique in microsomal fractions from vitamin E-deficient and control rats. The induction period that normally precedes the ascorbate/ADP/Fe3+-induced lipid peroxidation was taken as reflecting the microsomal vitamin E content and was found to be 5-6-fold decreased in microsomal fractions from vitamin E-deficient rats. Supplementation of microsomal fractions from vitamin E-deficient rats with exogenous vitamin E partially restores the induction period observed in that from control rats. The decrease in chemiluminescence intensity and the increase in the induction period both correlate linearly with the amount of vitamin E added. However, the efficiency of exogenous vitamin E is about 50-fold lower than that exerted by the naturally occurring vitamin E in microsomal membranes. These observations are discussed in terms of the process of re-incorporation of vitamin E into membranes, the experimental model for lipid peroxidation selected, and the method to evaluate lipid peroxidation, namely low-level chemiluminescence.

Animals↗
PubMed1984

A novel biologically active seleno-organic compound--I. Glutathione peroxidase-like activity in vitro and antioxidant capacity of PZ 51 (Ebselen).

a synthetic seleno-organic compound, 2-phenyl-1,2-benzoisoselenazol-3(2H)-one (PZ 51), exhibits GSH peroxidase-like activity in vitro, in contrast to its sulfur analog, PZ 25. In addition, PZ 51 behaves as an antioxidant shown by a temporary protection of rat liver microsomes against ascorbate/ADP-Fe-induced lipid peroxidation, an effect also elicited by PZ 25 but to a smaller extent. This protection against lipid peroxidation is independent of GSH and of P-450 monooxygenase activity.

Animals↗
PubMed1984

In situ behaviour of D(-)-lactate dehydrogenase from Escherichia coli.

Some kinetic properties of the D(-)-lactate dehydrogenase (EC 1.1.1.28) of Escherichia coli have been investigated. There were marked differences between the kinetic properties of the enzyme studied in situ compared with the in vitro D(-)-lactate dehydrogenase. D(-)-Lactate dehydrogenase in situ showed high substrate inhibition with pyruvate over the pH range 6.0-7.0, whereas the enzyme in vitro did not. The pH optimum for pyruvate reduction by the in situ D(-)-lactate dehydrogenase ranged between pH 7.5 and 7.8, whereas the in vitro enzyme showed its pH optimum between pH 6.8 and 7.0. The pK values of the prototropic groups that controlled the enzymatic activity shift to the acidic region for the in vitro enzyme with respect to the in situ enzyme. In vitro D(-)-lactate dehydrogenase exhibits homotropic interactions with its substrate, pyruvate and its coenzyme, NADH, at pH values ranging between pH 6.0 and 8.5, but the in situ enzyme showed homotropic interactions neither with pyruvate nor with NADH at all pH values studied.

Escherichia coli↗
PubMed1983

Antioxidant effect of diethyldithiocarbamate on microsomal lipid peroxidation assessed by low-level chemiluminescence and alkane production.

Different thiol-containing compounds, such as diethyldithiocarbamate (DDC), glutathione, penicillamine, and dithioerythritol have been chosen to study their effect on ascorbate/Fe-ADP-induced lipid peroxidation, detected by low-level chemiluminescence and alkane production. In the concentration range used, these thiols exerted a temporary protection against lipid peroxidation by lengthening the induction period; after overcoming this induction period, no substantial inhibition of either chemiluminescence or alkane production was observed. DDC was effective in protecting against lipid peroxidation in the nanomolar range, whereas the group of other thiol-containing molecules operated in the millimolar range.

Adenosine Diphosphate↗
PubMed1983

Effects of 4-hydroxynonenal on isolated hepatocytes. Studies on chemiluminescence response, alkane production and glutathione status.

The effect of 4-hydroxy-2,3-trans-nonenal, a diffusible product of lipid peroxidation, on isolated hepatocytes was evaluated with two non-invasive techniques measuring low-level chemiluminescence and alkane evolution. Oxygen-induced low-level chemiluminescence and ethane and n-pentane formation by hepatocytes is enhanced over 7-fold in the presence of 4-hydroxynonenal (2 mM). Glutathione-depleted hepatocytes show a higher increase than controls in both low-level chemiluminescence and alkane formation upon supplementation with 4-hydroxynonenal. The effects on both parameters are diminished by vitamin E pretreatment of rats and are absent under anaerobiosis. At variance with chemiluminescence and alkane formation, 4-hydroxynonenal does not elicit a concomitant increase in malonaldehyde or diene-conjugate formation. Addition of 4-hydroxynonenal to a suspension of hepatocytes causes a rapid loss of cellular glutathione in the form of a glutathione conjugate with the alkenal as observed with high-pressure liquid-chromatographic analysis. The reaction between glutathione and 4-hydroxynonenal proceeds also spontaneously in vitro at 1:1 stoichiometry. The cellular effects of 4-hydroxynonenal evaluated by low-level chemiluminescence and alkane formation are independent of the formation of a glutathione conjugate and seem to rely on the remaining not-bound 4-hydroxynonenal. The sensitivity of 4-hydroxynonenal-enhanced chemiluminescence and alkane formation to free-radical quenchers suggests the participation of a free-radical propagation process.

Aldehydes↗
PubMed1983

Singlet oxygen formation detected by low-level chemiluminescence during enzymatic reduction of prostaglandin G2 to H2.

Addition of arachidonic acid to a suspension of ram vesicular gland microsomes or purified prostaglandin synthase, causes a rapid burst of light emission in the range 600-750 nm, as detected by single-photon counting. Maximal light emission intensity is obtained within 15-30 s after the addition of arachidonic acid and is followed by a rapid decay to the background level. The intensity of chemiluminescence is dependent on the amount of ram vesicular gland microsomes or isolated prostaglandin synthase and arachidonic acid concentration (Km about 6 microM). Spectral analysis of arachidonic acid-induced photoemission of isolated prostaglandin synthase in the range 600-750 nm showed two distinctive peaks at about 634 and 703 nm. The similar relative intensities of these peaks, along with the lower intensity at about 668 nm is indicative of singlet oxygen dimol emission. Chemiluminescence with arachidonate is enhanced by 1,4-diazabicyclo[2,2,2]octane and inhibited by azide, indomethacin, acetylsalicylic acid and beta-carotene. Cooxygenation substrates such as phenol, hydroquinone and reduced glutathione, inhibited the arachidonic acid-induced chemiluminescence. Dioxygen is a requirement for the observation of singlet oxygen dimol emission with arachidonic acid as a substrate for ram vesicular gland microsomes or purified prostaglandin synthase. However, when prostaglandin G2 is substituted for arachidonic acid, light emission is not dependent on oxygen. Thus, singlet oxygen can be formed in the dismutation reaction, 2 PGG2 leads to 2 PGH2 + 1O2, catalysed by prostaglandin hydroperoxidase.

Animals↗
PubMed1983

Evidence for lipid peroxidation by paraquat in the perfused rat lung.

In order to evaluate the effect of paraquat on oxidative radical reactions in the lung, we studied MDA production and chemiluminescence (spontaneous and tBuOOH-induced) in the isolated rat lung. After 2 hr of perfusion with 3.0 mM paraquat, MDA content in lung homogenates was 16 +/- 7 nmol/gm dry weight higher than in control lungs (mean +/- S.E., n = 7, p less than 0.05 by paired test); during 30 min of perfusion, malondialdehyde efflux was 33 +/- 15 nmol/gm dry weight higher than in control perfusates (n = 6, p less than 0.05). Spontaneous chemiluminescence was not augmented by 2 hr of perfusion with concentrations of paraquat ranging from 0.75 to 6.0 mM. On the other hand, tBuOOH-induced chemiluminescence was 17% +/- 3 higher immediately after the addition of hydroperoxide and reached a 16% +/- 6 higher plateau for the paraquat-perfused lungs than for control lungs (n = 10, p less than 0.05). Spectral analysis of the light emitted during induced chemiluminescence demonstrated peak intensity between 630 and 730 nm for both control and paraquat-treated lungs. Increased MDA production and increased induced chemiluminescence indicate that perfusion with paraquat enhances lipid peroxidation in the isolated rat lung.

Animals↗
PubMed1982

Lactate dehydrogenase in Phycomyces blakesleeanus.

1. An NAD-specific L(+)-lactate dehydrogenase (EC 1.1.1.27) from the mycelium of Phycomyces blakesleeanus N.R.R.L. 1555 (-) was purified approximately 700-fold. The enzyme has a molecular weight of 135,000-140,000. The purified enzyme gave a single, catalytically active, protein band after polyacrylamide-gel electrophoresis. It shows optimum activity between pH 6.7 and 7.5. 2. The Phycomyces blakesleeanus lactate dehydrogenase exhibits homotropic interactions with its substrate, pyruvate, and its coenzyme, NADH, at pH 7.5, indicating the existence of multiple binding sites in the enzyme for these ligands. 3. At pH 6.0, the enzyme shows high substrate inhibition by pyruvate. 3-hydroxypyruvate and 2-oxovalerate exhibit an analogous effect, whereas glyoxylate does not, when tested as substrates at the same pH. 4. At pH 7.5, ATP, which inhibits the enzyme, acts competitively with NADH and pyruvate, whereas at pH 6.0 and low concentrations of ATP it behaves in a allosteric manner as inhibitor with respect to NADH, GTP, however, has no effect under the same experimental conditions. 5. Partially purified enzyme from sporangiophores behaves in entirely similar kinetic manner as the one exhibited by the enzyme from mycelium.

Adenosine Triphosphate↗
PubMed1982

Influence of pH on the allosteric properties of lactate dehydrogenase activity of Phycomyces blakesleeanus.

1. Lactate dehydrogenase from mycelium of Phycomyces blakesleeanus showed positive homotropic interactions with NADH at all pH values studied (pH 5.0-7.7). The calculated values for the first and last intrinsic association constants remained unaltered with pH, in contrast with the Hill coefficient value, which varied significantly, reaching its maximum values at pH 6.0 and 7.7. This suggests the hypothesis that pH regulates these homotropic effects by changes in the value of the intermediate intrinsic association constants. 2. From pH 7.2 to 7.7 lactate dehydrogenase exhibited, likewise, positive homotropic interactions with pyruvate. There were practically no changes in the first and last intrinsic association constants and in Hill coefficient values with pH. At pH values below 7.2 (pH 5.0-6.8) the enzyme showed high substrate inhibition, which was highly dependent on pH, NADH concentration and temperature. By way of substrate inhibition pH regulates, primarily, lactate dehydrogenase activity towards pyruvate, since the homotropic effects appear not to be dependent on pH. 3. Fructose 1,6-bisphosphate is a true allosteric effector of lactate dehydrogenase of Phycomyces blakesleeanus. it decreases positive co-operativity with NADH, and on the other hand pyruvate co-operativity turns into mixed co-operativity. In addition, the effector decreases the inhibitory effect caused by pyruvate.

Allosteric Site↗
PubMed1982

Active oxygen metabolites and their action in the hepatocyte. Studies on chemiluminescence responses and alkane production.

"Oxidative stress" takes place in animal tissues when the balance between the cellular defense mechanisms (glutathione cycle, superoxide dismutase, catalase, vitamin E, etc.) and conditions capable of triggering oxidative reactions is altered. The oxidative reactions which occur under a variety of conditions were assessed by two non-invasive methods, low-level chemiluminescence and volatile hydrocarbon production. Oxidative stress induced by hyperoxia or organic hydroperoxides in isolated hepatocytes or the perfused liver, respectively, is accompanied by low-level chemiluminescence, the intensity of which is enhanced upon perturbation of the glutathione cycle system, i.e., glutathione depletion and/or selenium deficiency. Oxidative stress during redox cycling of paraquat, when infused into the perfused liver, is not accompanied by light emission, whereas menadione, a substance also capable of redox cycling, was found to elicit photoemission under similar conditions. The basal rates of ethane release by the perfused liver are enhanced during oxidative conditions such as metabolism of hydroperoxides, paraquat redox cycling, and ethanol oxidation. Alkane release during the latter involves the participation of alcohol dehydrogenase and further products of ethanol oxidation, i.e., acetaldehyde, as well as free radicals in some stage of the process. In vivo ethane release by animals with adjuvant arthritis was found higher than in controls, presumably due to a systemic response of liver to inflammation.

Alkanes↗