GROWTH, DEVELOPMENT AND RESPIRATION IN THE OVULES OF ZEPHYRANTHES LANCASTERI AT DIFFERENT STAGES OF MATURATION

Changes in the rate of respiration and activity of succinicdehydrogenase in the ovules, embryo and endosperm have beeninvestigated at different stages of seed development in Z. lancasteri.An attempt has been made to find any possible relation betweenphysiological changes and the growth pattern of ovule. The fertilizedovules mature into seeds in 16–19 days from the time ofpollination. The respiratory course and the growth of ovulesfollow identical trends. The curve for oxygen uptake is characterizedby at least two peaks; the first on the sixth or seventh dayis related to the laying down of walls in the coenocytic micropylarchamber of the endosperm and the second on the 11th day precedesthe elongation of cotyledon. In excised endosperm the activityof succinic dehydrogenase is highest during its transformationfrom freenuclear to cellular state, while in the embryo it isso after the attainment of mature size. ¹Present address: AEC Plant Research Laboratory, Michigan StateUniversity, East Lansing, Mich., U.S.A.     

Structure, Composition and Physiological State of the Endosperm of Phoenix dactylifera L

The date endosperm consists of living cells with the same generalcellular structure throughout the seed. The major storage products,as shown by histochemical staining, are lipid, stored as numeroussmall lipid bodies which fill the cytoplasm, and protein, aslarge but variably-sized protein bodies. Nuclei are presentbut lack large amounts of heterochromatin. Plastids and mitochondriaare present but infrequently seen and have poorly developedinternal membranes. No endoplasmic reticulum or dictyosomesare present before or after hydration. The cell wall is thickexcept in areas of pit fields and consists of three layers whichdiffer in their staining behaviour: middle lamella, thickenedwall and inner wall. Both the endosperm and embryo of the imbibedseed undergo aerobic respiration, but the embryo respires ata more rapid rate than does the endosperm. A small area of theendosperm around the distal pole of the cotyledon shows histochemicallydetectable levels of succinic dehydrogenase. Phoenix dactylifera L, date palm, endosperm, ultrastructure, histochemistry, respiration, succinic dehydrogenase     

High Ambient Carbon-Dioxide does not Affect Respiration by Suppressing the Alternative, Cyanide-resistant Pathway

Total dark respiration (v_t), cytochrome pathway (v_eyt), alternativepathway (v_alt) and residual (v_res) respiration were measuredin Lemna gibba plants, by the use of pathway inhibitors. NaCNwas used to inhibit v_eyt and SHAM (salicylhydroxamic acid) toinhibit v_alt. Residual respiration (v_res) was about 5% of v_t.The effect of high (100 Pa) and low (0 Pa) carbon dioxide partialpressure ([CO₂]) on v_t, v_eyt and v_alt was determined from bothCO₂ efflux and O₂ uptake measurements. The higher [CO₂] suppressedv_t by about 30%. When respiration operated through the cytochromepathway only, in the absence of v_alt, it was suppressed by about12% as measured by the O₂ uptake of submerged Lemna fronds orby about 40% as measured by CO₂ efflux from floating fronds.The higher [CO₂] treatment had only a small effect on respiration,when v_alt alone operated. There was no evidence of a specificsuppression of the v_alt pathway by high [CO₂]. Succinic dehydrogenaseactivity of the mitochondria of roots of Medicago sativum wasreduced by 18%, when the mitochondria were pre-treated with120 as compared to 34 Pa [CO₂]. There was no such effect oncytochrome c oxidase activity of mitochondria under the sameconditions. It is concluded that there is no evidence for the hypothesisthat the high [CO₂] suppression of respiration is a result ofa CO₂ effect on the non-phosphorylating alternative respirationpathway.Copyright 1995, 1999 Academic Press Lemna gibba, Medicago sativum, respiration, cytochrome pathway, alternative pathway, cyanide-insensitive pathway, carbon dioxide, succinic dehydrogenase, cytochrome c oxidase     

A Comparison of Leek (Allium porrum) and Onion (Allium cepa) Seed Development

Leek and onion seed dry weight increased exponentially for thefirst 31 days after flowering (DAF) but thereafter the increasein dry weight was slower. Before maximum seed dry weight wasreached at 45 DAF in onion and 59 to 66 DAF in leek, seed moisturecontent, seed oxygen uptake and conductivity of the seed steepwater fell from initially high levels. Although some seeds germinated31 DAF in both species, full germination in both was not reacheduntil 66–80 DAF. Tolerance of the seed to artificial dryingimmediately after harvesting occurred 45 DAF in onion and 74DAF in leek. Free nuclear division continued in the endospermuntil 17–22 DAF in onion and until 31–35 DAF inleek but it was not until 45 DAF in onion and 66 DAF in leekthat the embryo and endosperm filled the cavity formed by thepericarp. After formation of cell walls in the endosperm thepattern of change in cell number in both species was similar.The shrunken appearance of the seed coat in leek, which occurredearly in seed development, was associated with the period offree nuclear division in the endosperm and, in addition, thepericarp was thinner than in onion. There was no evidence thatthe shrunken seed coat early in development was associated withself as opposed to open-pollination. Allium porrum, Allium cepa, seed development, endosperm, embryo, cell number, germination, respiration, seed leachates     

The physical and chemical properties of a chicken egg white glycoprotein purified by nondenaturing methodology

The oxidation of ethanol by the liver produces acetaldehyde, which is a highly reactive compound. Low concentrations of acetaldehyde inhibited mitochondrial respiration with glutamate, β-hydroxybutyrate, or α-ketoglutarate as substrates, but not with succinate or ascorbate. High concentrations led to respiratory inhibition with all substrates. Inhibition of succinate- and ascorbate-linked oxidation by acetaldehyde correlates with the inhibition of the activities of succinic dehydrogenase and cytochrome oxidase. A site more sensitive to acetaldehyde appears to be localized prior to the NADH-ubiquinone oxidoreductase segment of the respiratory chain. Acetaldehyde inhibits energy production by the mitochondria, as evidenced by its inhibition of respiratory control, oxidative phosphorylation, the rate of phosphorylation, and the ATP-³²P exchange reaction. Energy utilization is also inhibited, in view of the decrease in both substrate- and ATP-supported Ca²⁺ uptake, and the reduction in Ca²⁺-stimulated oxygen uptake and ATPase activity. The malate-aspartate, α-glycerophosphate, and fatty acid shuttles for the transfer of reducing equivalents, and oxidation by mitochondria, were highly sensitive to acetaldehyde. Acetaldehyde also inhibited the uptake of anions which participate in the shuttles. The inhibition of the shuttles is apparently caused by interference with NAD⁺-dependent state 3 respiration and anion entry and efflux. Ethanol (6–80 mm) had no significant effect on oxygen consumption, anion uptake, or mitochondrial energy production and utilization. The data suggest that acetaldehyde may be implicated in some of the toxic effects caused by chronic ethanol consumption.     

Effect of Some Dithiocarbamates on Respiration Activity of a Myxomycete

The presenl article aims at giving some information concerning the mode of action of three dithiocarbamates: ferric dimethyldithiocarbamate (Ferbam), zinc dimethyl-dithiocarbomate (Ziram) and tetramethylthiuram disulfide (Thiram). These inhibitors were tested on the respiratory activity of the Myxomycete: Physarum polycephalum. Its development is strongly inhibited at 285 mg/l. With such concentration, respiration measurements confirmed total inhibition of O₂ uptake. After isolation of the mitochondria, the tests showed that there may be an inhibition of the succinic oxidase activity of this particulate fraction, at 28.5 mg/l of Thiram. Heavy concentration of reduced glutathione (1.4 %) reversed partially this Inhibition. This seems to prove that succinic dehydrogenase is a typical SH-type enzyme.     

Effetto di Inibitori Della Sintesi Proteica Sull'aumento di Attività Enzimatiche e sul Risveglio del Metabolismo Nell'Endosperma di Semi di Ricino in Germinazione

Abstract

Effects of inhibitors of protein synthesis on the development of metabolic activity in the endosperm during the germination of castor bean seeds. — The effect of chloramphenicol, streptomycin and actinomycin-C on the increase of the activities of glyceroaldehyde-phosphate dehydrogenase, aldolase, glucose-6-phosphate dehydrogenase, fructose 1–6 diphosphate-1-phosphatase, phosphomonoesterase, in the endosperm of germinating castor bean seeds was investigated.

In all cases, the protein synthesis inhibitors depressed the activation of the enzymes tested: in particular, actinomycin (50 μg/ml) completely suppressed the increase of the activities.

The development of the rate of oxygen uptake and the conversion of fats to sugars was strongly affected by the inhibitors.

These data suggest that the increase of the activities of several enzymes in the germinating endosperm is dependent on enzyme synthesis rather than on the conversion from the inactive to the active form of the enzymes.     

Intermediate Energy Metabolism of Leptospira

Metabolic studies were performed on three representative serotypes of Leptospira: a water isolate designated B(16) and two pathogenic serotypes, pomona and schueffneri. Examination of whole cells of B(16) for their ability to oxidize various substrates revealed that oleate significantly stimulated oxygen uptake. The respiratory quotient of 0.7 implied that oleate was degraded to carbon dioxide and water. Other substrates, such as carbohydrates, alcohols, intermediates of the citric acid cycle, and short-chain acids, including selected amino acids, did not stimulate endogenous respiration of whole cells. No oxygen uptake could be measured when cell-free extracts were tested with the substrates used with whole cells. Enzymatic analyses of cell-free extracts of the three strains demonstrated enzymes of the citric acid cycle, enzymes of the glycolytic and pentose pathways, and the general acyl coenzyme A dehydrogenase required for beta-oxidation of fatty acids. Strain B(16) and the two pathogenic serotypes appeared to possess similar metabolic capabilities. Enzymatic data might also explain the apparent inability of B(16) to oxidize other substrates; kinases necessary for activation of common nonphosphorylated compounds were not detected in leptospiral extracts. These findings emphasized the dependence of leptospiral growth upon long-chain fatty acids.     

Effects of phthalate esters on the respiration of rat liver mitochondria.

The effects of phthalate esters on the oxidation of succinate, glutamate, beta-hydroxybutyrate and NADH by rat liver mitochondria were examined and it was found that di-n-butyl phthalate (DBP) strongly inhibited the succinate oxidation by intact and sonicated rat mitochondria, but did not inhibit the State 4 respiration with NAD-linked substrates such as glutamate and beta-hydroxybutyrate. However, oxygen uptake accelerated by the presence of ADP and substrate (State 3) was inhibited and the rate of oxygen uptake decreased to that without ADP (State 4). It was concluded that phthalate esters were electron and energy transport inhibitors but not uncouplers. Phthalate esters also inhibited NADH oxidation by sonicated mitochondria. The degree of inhibition depended on the carbon number of alkyl groups of phthalate esters, and DBP was the most potent inhibitor of respiration. The activity of purified beef liver glutamate dehydrogenase [EC 1.4.1.3] was slightly inhibited by phthalate esters.     

Oxidative Activity of Particles prepared from Lettuce Seedlings

A particulate fraction similar to mitochondria was isolatedfrom germinating lettuce seedlings. The endogenous oxygen uptake of this fraction was very high.It could oxidize most of the Krebs cycle intermediates, theoxidation of l-malate being particularly marked. Glutamic, glycollic, and formic acids could also be oxidizedto a certain extent. Fatty acids inhibited the endogenous respiration,the inhibition increasing with increased concentration. Coumarinand thiouren do not seem to have any direct effect on the respiratoryenzyme systems. They apparently affect the respiration of wholeseeds only indirectly, maybe through some germination regulatingmechanism not yet known.     

A New Procedure for the Calculation of Oxygen Diffusion Resistance in Legume Nodules from Flow-through Gas Analysis Data

Plants of white lupin (Lupinus albus L cv Multolupa) and soyabean(Glycine max L cv Clarke) were grown in controlled-environmentcabinets, subjected to various stresses and their nodular nitrogenaseactivity and total root respiration measured When these measurementswere used to calculate nodular oxygen diffusion resistance,using a simplified equation for Fick's first law of diffusion,it was found that the apparent resistance of stressed nodulesincreased anomalously with decreases in external oxygen concentrationA new analysis procedure is proposed to alleviate this anomalyThis procedure also uses the simplified Fick's law equationbut includes a respiratory contribution to the total oxygenflux across the diffusion barrier which is not coupled to nitrogenaseactivity Also, resistance is modelled as an exponential functionof external oxygen concentration Use of this analysis procedureproduces realistic values for total resistance and providesa characterisation of this resistance into a minimum value andan adjustment factor for changes in external oxygen It is postulatedthat the additional respiration component represents the activityof nodule cortex cells involved in the diffusion barrier, particularlythat of vascular bundles Oxygen diffusion resistance, nodule, nitrogen fixation, respiration     

Metabolism of Slices of the Tomato Stem

The rate of respiration of tomato stem slices varied considerably,the highest values (Qo₂, 2–3) being obtained for plantsin a good nutritional state, and the lowest (Qo₂, 1) in starvedplants. The respiratory quotient of 1.0 remained constant. Glucose fermentation was found to follow both glycolytic andalcohol fermentation pathways, the ratio of ethyl alcohol: lacticacid being 6.6:1. Fermentation seems to take place accordingto the Embden-Meyerhof scheme, as shown by the presence of someof these enzymes operative in this scheme and by inhibitionexperiments. In the presence of oxygen there was no formationof alcohol or lactic acid. Pyruvate added to tomato stem slices was metabolized by directoxidation to acetic acid and by dismutation to lactic and aceticacids and CO₂ The metabolism of acetic acid was demonstratedby its condensation with oxaloacetic acid to form citrate, thisbeing the second time that synthesis of citric acid by thismechanism has been found in plants. The presence of aconitase,of isocitric dehydrogenase, of succinic dehydrogenase, and ofmalic dehydrogenase, as well as the inhibition of respirationby malonic acid, favour the hypothesis that oxidation of carbohydratein tomato stem slices proceeds via the citric acid cycle. Thepossibility of an auxiliary route, the malic acid oxidationpathway, also was demonstrated. Tomato stem tissue anaerobicallysplit malic acid into glycolic acid. The further oxidation ofglycolic, glyoxylic, and formic acid was demonstrated. In experiments with C¹⁴-labelled acetate and butyrate a dilutionof the C¹⁴-labelled acids was found after incubation indicatingnew formation of these acids and of active participation offatty acid metabolism in the metabolic activities of the tissue. With the exception of alanine, added amino acids produced adefinite increase in O₂ uptake without extra formation of ammonia. Experimental demonstration of the possibility of electron transportfrom substrate to molecular oxygen in respiration via polyphenoloridase was provided by the attainment in a tomato tissue homogenateof a coupled oxidation-reduction between -ketoglutarate andcatechol with DPN and tyrosinase as the catalysts. The presenceof cytochrome oxidase was also demonstrated. Thus both systemspossibly may take part in the respiration of tomato stem slices.     

The Respiration Rate of Tomato Stem Tissue Infected by Tomato Aucuba Mosaic Virus

The oxygen uptake of stem tissue slices of Lycopersicum esculentumvar. ‘Stonor's Moneymaker’, in 0·035 M. Sorensonphosphate buffer was measured manometrically; the optimum pHwas found to be 5·2. With tomato-stem tissue systemicallyinfected by virus (tomato aucuba mosaic), the respiration ratewas always below that of healthy tissue when equal numbers ofrespiratory systems were compared. When the respiration ratewas expressed in terms of oxygen uptake per unit fresh or dryweight, the respiration rate of the virus-infected tissue couldbe greater or less than that of the healthy tissue dependingon the growth conditions of the experimental material. Underconditions favouring the complete development of disease symptoms,virus infection decreased the fresh and dry weight of the cell.For this reason fresh weight and dry weight were not consideredsuitable criteria to express the respiration rates as theseunits contained different numbers of respiratory systems inhealthy and virus-infected tissue.     

Respiration and dehydrogenase activity of pigmentless variants of Staphylococcus aureus]

Respiration and dehydrogenase activity were studied in four pigmentless mutants of Staphylococcus aureus 209-P. The rate of oxygen uptake from the incubation medium was higher in the mutants than in the parent strain. The rate of respiration and the activity of dehydrogenase in the pigmentless mutants were found to depend on the permeability of the cell membrane for substrates of respiration; carotenoids are presumed to be involved in this permeability.     

Respiration of Malate and Glutamate in Symbiotic Frankia Prepared from Alnus incana

Frankia vesicle clusters were prepared from root nodules ofAlnus incana (L.) Moench inoculated either with a local sourceof Frankia or with Frankia Cpll. The capacity of vesicle clustersto respire was investigated by respirometric and enzymologicalstudies. Simultaneous addition of malate, glutamate, and NAD⁺supported respiration in both types of Frankia, though at asmaller rate compared to the substrates NADH or 6-phosphogluconate.The saturating concentrations of malate and glutamate were alsomuch higher than with the other substrates. No respiration wassupported by succinate. Activity of the enzymes malate dehydrogenase(EC 1.1.1.37 [EC] ) and glutamate oxaloacetate transaminase (EC 2.6.1.1 [EC] )was demonstrated in crude extracts from both types of symbioticFrankia. Their maximum rates were high enough to account forthe respiration of malate and glutamate. This respiration wasinhibited by mersalylic acid, an inhibitor of the dicarboxylateshuttle in mitochondria, but it was shown that inhibition ofrespiration could be due to a direct effect on the enzymes.We conclude that respiration of malate and glutamate is mostlikely mediated by malate dehydrogenase and glutamate oxaloacetatetransaminase, but no explicit evidence for or against the presenceof a dicarboxylate carrier was found. The utilization of respiratorysubstrates was largely similar in the two types of Frankia,except for some differences in maximum rates and cofactor dependency. Key words: Actinorhizal symbioses, Alnus, dicarboxylate shuttle, Frankia, reducing power, respiration     

Endogenous and endobiotic induced reactive oxygen species formation by isolated hepatocytes.

The rat hepatocyte catalyzed oxidation of 2',7'-dichlorofluorescin to form the fluorescent 2,7'-dichlorofluorescein was used to measure endogenous and xenobiotic-induced reactive oxygen species (ROS) formation by intact isolated rat hepatocytes. Various oxidase substrates and inhibitors were then used to identify the intracellular oxidases responsible. Endogenous ROS formation was markedly increased in catalase-inhibited or GSH-depleted hepatocytes, and was inhibited by ROS scavengers or desferoxamine. Endogenous ROS formation was also inhibited by cytochrome P450 inhibitors, but was not affected by oxypurinol, a xanthine oxidase inhibitor, or phenelzine, a monoamine oxidase inhibitor. Mitochondrial respiratory chain inhibitors or hypoxia, on the other hand, markedly increased ROS formation before cytotoxicity ensued. Furthermore, uncouplers of oxidative phosphorylation inhibited endogenous ROS formation. This suggests endogenous ROS formation can largely be attributed to oxygen reduction by reduced mitochondrial electron transport components and reduced cytochrome P450 isozymes. Addition of monoamine oxidase substrates increased antimycin A-resistant respiration and ROS formation before cytotoxicity ensued. Addition of peroxisomal substrates also increased antimycin A-resistant respiration but they were less effective at inducing ROS formation and were not cytotoxic. However, peroxisomal substrates readily induced ROS formation and were cytotoxic towards catalase-inhibited hepatocytes, which suggests that peroxisomal catalase removes endogenous H(2)O(2) formed in the peroxisomes. Hepatocyte catalyzed dichlorofluorescin oxidation induced by oxidase substrates, e.g., benzylamine, was correlated with the cytotoxicity induced in catalase-inhibited hepatocytes.     

The Effect of pH on the Action of Respiratory Inhibitors in Avena fatua Seeds

The effects of pH on the action of sodium azide, a cytochome-oxidaseinhibitor, and salicylhydroxamate (SHAM), an alternative respirationinhibitor, on the respiration of dormant seeds of wild oat (Avenafatua L.; line AN-51) were studied. While pH had little effecton seed respiration in controls, it strongly affected the activityof azide. One mM azide inhibited seed respiration at pH5, butstimulated it at pH 7. SHAM (10 mM) completely inhibited thestimulation of respiration by 1 mM azide in an unbuffered medium,but failed to do so when the medium was buffered at pH 7. Inunbuffered medium, 10 mM SHAM actually augmented the stimulationof respiration by 0.25 mM azide to the same degree as when theazide solution was acidified to mimic the pH change incurredwith dissolution of 10 mM SHAM. These results suggest that theinhibitory effect of SHAM on the action of azide in an unbufferedsystem may in part be due to its acidification of the incubationmedium rather than by the inhibition of alternative oxidase.Lower pH favours the formation of the undissociated hydrazoatemolecules causing greater inhibition of cytochrome-oxidase andother azide-sensitive enzymes. Avena fatua L, wild oat, seed dormancy, azide, salicylhydroxamate     

Substrate Dependent Reversibility of Inhibition by β-Thujaplicin of Glucose and Acetate Respiration in Saccharomyces cerevisiae

Attempts to determine the rate of uptake of β-thujaplicin (TH) and its 1:1 rupric chelate (CuT⁺) in yeast cells showed in both cases a very rapid equilibration. In the concentration range where respiration is inhibited, most of the TH added was still present in the extracellular fluid, while CuT⁺ was almost completely removed. The inhibition of glucose respiration by TH was Completely removed when the cells were repeatedly washed, while inhibition of acetate respiration remained. Washing of CuT⁺ inhibited cells did not release the inhibition of either glucose or acetate respiration. The succinic acid dehydrogenase activity was the same in homogenates of both CuT⁺ and TH inhibited cells as in homogenates of non-inhibited cells. Prior to homogenization acetate respiration was inhibited by 60 to 70 per cent by both inhibitors. Alcohol dehydrogenase activity was likewise not affected by TH and CuT⁺. The formation of acetyl coenzyme A was inhibited by both TH and CuT⁺ to about the same extent as the respiration of acetate. It is concluded that TH inhibits the acetate respiration by inhibiting the formation of acetyl coenzyme A from acetate. The same site of action is plausible also for CuT⁺ but in that case, additional sites of inhibition may also be located in the reactions in the Krebs cycle.     

轻度水分胁迫的小麦幼苗中与呼吸有关的几种酶活性变化

轻度水分胁迫使小麦幼苗叶片呼吸升高时,叶中琥珀酸去氢酶和细胞色素氧化酶活性均明显升高;而同样胁迫使根呼吸下降时,根中这两种酶活性均明显下降。叶和根中ATP酶分解活性在胁迫下都明显升高。轻度水分胁迫使叶片过氧化氢酶活性升高。叶中有明显的乙醇酸氧化酶活性,抗旱品种的酶活性较高,胁迫使此酶活性降低。     

Variation in Photorespiration in Lolium

The rate of photorespiration in several grass species was shownto be highly variable and dependent on the species, genotype,and conditions under which the plants were grown. Photorespiration,measured as oxygen uptake, was negligible in Cenchrus ciliarisand Paspalum dilatatum but significant in Lolium spp. and Festucaarundinacea. There were significant differences in the rateof photorespiration among plants within a Lolium populationof diverse origin and these differences were independent ofthe conditions under which the plants were grown. Among thetemperate grasses there was a significant correlation betweenphotorespiration and the CO₂-compensation concentration andboth parameters were very low in P. dilatatum. Plants grownin day/night temperatures of 15/10 °C compared with 25/20°C had faster rates of dark respiration but slower ratesof light respiration when measured at the same temperature.Photorespiration was faster than dark respiration although differencesin respiration among plants in the light were not shown in thedark.