Unlabeled CO2 Research Articles

Partitioning of carbon in split root systems of barley: effect of temperature of the root

The time-course of partitioning of recently fixed photosynthate to a barley root was followed during a step change in the temperature of the root. The induced change in partitioning had a response time of about 1 h, whilst the change in respiratory loss from the root of labelled and unlabelled CO was much faster. These observations suggest that while a sudden temperature change has an immediate effect upon the metabolic rate of the roots it is not the metabolic rate as such which influences import (sink strength). The reported temperature responses are consistent with the supposition that the size of the cytosolic pool of sugar in the root, which is supplied directly by phloem import and used as a substrate for root metabolism, appears to control import into the root.

The Energetics of Reproduction in the Common Shrew (Sorex araneus): A Comparison of Indirect Calorimetry and the Doubly Labeled Water Method

Energy expenditure of the common shrew (Sorex araneus) was measured by two techniques, indirect calorimetry (IC) and the doubly labeled water (DLW) method. Daily energy expenditure (DEE) did not increase significantly during pregnancy when measured by IC (days 0-7, 56.5 kJ · d⁻¹; days 8-14, 60.8 kJ · d⁻¹; days 15-20, 65.0 kJ · d⁻¹) or when measured by DLW (days 0-7, 70.1 kJ · d⁻¹; days 8-14, 72.6 kJ · d⁻¹; days 15-20, 74.9 kJ · d⁻¹). Food intake also did not increase significantly during pregnancy. We hypothesize that there was a decrease in a component of the energy budget not directly associated with reproduction, termed metabolic compensation. The energetic cost of lactation was far greater than the cost of pregnancy. After the initial increase the maternal DEE, measured by DLW, remained remarkably constant throughout lactation (days 08, 154.3 kJ · d⁻¹; days 9-16, 158.9 kJ · d⁻¹; days 17-23, 181.5 kJ · d⁻¹), although food intake continued to increase. There may be further metabolic compensation during lactation. During both pregnancy and lactation there was a discrepancy between measurements of energy expenditure obtained by the two techniques. In pregnancy the higher DLW expenditures were attributed to a difference in environmental conditions. During lactation we conclude that the high DLW energy expenditures may have in part been due to an exchange of unlabeled CO₂ from the litter to the mother.

Retrobiosynthetic analysis of carbon fixation in the phototrophic eubacterium Chloroflexus aurantiacus

The phototrophic bacterium Chloroflexus aurantiacus does not use any of the known autotrophic CO2 fixation pathways. There is evidence for a new cyclic autotrophic pathway in which acetyl-CoA is converted to 3-hydroxypropionate and further to succinate and malate. This hypothesis was tested by feeding growing cultures during several generations with 3-hydroxy[1-13C]propionate, [1-13C]acetate, or [2-13C]acetate, in addition to unlabeled CO2. The relative 13C content of individual carbon atoms in biosynthetic amino acids and nucleosides was determined by 1H- and 13C-NMR spectroscopy. 13C coupling patterns were analyzed by two-dimensional 13C-TOCSY experiments which were optimized for the analysis of multiply 13C-labeled biosynthetic samples. From the 13C enrichments of amino acids and nucleosides, the labeling patterns of central metabolic intermediates were evaluated by a retrobiosynthetic approach. Both 3-hydroxypropionate and acetate were incorporated into all central metabolic pools. The 13C labeling and coupling patterns suggest a novel carbon fixation pathway via 3-hydroxypropionate. Specifically, we propose that acetyl-CoA is carboxylated to malonyl-CoA which is reduced under formation of 3-hydroxypropionyl-CoA. Dehydration and reduction yield propionyl-CoA which is converted to succinate by a second carboxylation reaction. The net product of autotrophic carbon fixation appears to be glyoxylate. However, it is not yet known how glyoxylate is channeled into anabolic metabolism. Assimilation of acetate can proceed via the CO2 fixation pathway, but also via the glyoxylate pathway.

Isotopic dilution of CO2 as an estimate of CO2 production during substrate oxidation studies

The rate of oxidation of a substrate is often divided by the numerical estimate of a correction factor for the proportion of labeled CO2 that is excreted during the course of the experiment. This factor, derived from tracer studies of bicarbonate kinetics, equals the product of the net rate of respiratory CO2 excretion and the ratio of the plateau isotopic enrichment of CO2 to the rate of infusion of labeled bicarbonate. The inverse of this ratio may be equated to the rate of appearance of unlabeled CO2. When one substitutes the expression for the correction factor into a typical equation for the rate of substrate oxidation, the rate of appearance of CO2 substitutes for both net CO2 excretion and the correction factor in the original oxidation equation. Thus the rate of appearance of CO2 is an index of CO2 production that takes into account both net fixation of CO2 and isotopic exchange of labeled CO2; it also may have application as an index of net CO2 production.

Adsorption state conversion of CO on Fe(100)

The conversion of adsorbed CO among different adsorption states on an Fe(100) surface was investigated through the coadsorption of labelled 13CO and unlabelled CO. In these experiments selected adsorption states were initially populated by one type of CO and the surface was then exposed to the other CO adsorbate. While no conversion occurred between the β-CO states and the α states, or between the α 2 and α 1 states, significant conversion was observed between the α 2 and α 3 states. A relative energy profile of the adsorption state conversions is proposed based on these observations.

An estimation of microbial death rate and limitations of N or C during wheat straw decomposition

The effect of initial microbial biomass size on subsequent straw decomposition was evaluated using labeled wheat straw ( Triticum aestivum L.) which had been leached to remove the readily-available C fraction. The leached, labeled straw was amended with 0.5,10 or 15% (w/w) unlabeled glucose to grow an unlabeled microbial biomass derived from readily-available C during incubation at 23° C. The glucose amendments had no effect on straw decomposition as measured by 14CO 2-C evolution, indicating that the microbial biomass derived from glucose was unable to utilize the nonsoluble portion of straw. Based on the evolution of unlabeled CO 2-C, the glucose-derived microbial biomass was estimated to have a half-life of 5 days with 55% of the C in dead microbial biomass forming new microbial material. The C and N pathways through two microbial pools are postulated to account for the lack of 14CO 2-C evolution in response to glucose amendment and for observed concurrent N immobilization and mineralization. The microbial biomass derived from the readily-available C (biomass pool A) rapidly consumes the available C and dies. Meanwhile, a different microbial biomass is growing on the less readily-available C components of straw (biomass pool B). Its growth is slower but is sustained for longer. The N mineralized from dead biomass pool A is immobilized into the biomass pool B resulting in net immobilization of N. This concept explains the commonly-observed concurrent mineralization and immobilization of N during periods of net immobilization. Evidence of two microbial pools, one C limited and one N limited, was supported by increased CO 2-C evolution following the addition of either glucose or kno 3 after 10 days to straw decomposing in the absence of added mineral N. Furthermore, the addition of glucose at 10 days did not influence the evolution of 14CO 2-C from 14C-labeled straw with or without the addition of N at the start of the experiment.

Mechanistic studies on the carbonylation of amidohydrocarbylnickel derivatives

The amidohydrocarbylnickel(II) complexes Ni(R)[N(SiMe 2CH 2PR′ 2) 2] (R′  Ph, R  CH 3, CHCH 2, C 6H 5, and C 6H 4- p-NMe 2; R′  Me, R  C 6H 5) react with carbon monoxide to generate nickel(0) derivatives of the formula Ni(CO) 2-[R CN(SiMe 2CH 2PR′ 2)O SiMe 2CH 2PR′ 2]. An intermediate in this process is the acylnickel species Ni(COR)[N(SiMe 2CH 2PR′ 2) 2] which has been isolated for all but the vinylnickel derivative; in this last case, addition of one equiv. of CO generates a 50% conversion to Ni(CO)[η 2-CH 2CH) CN(SiMe 2CH 2PPh 2)O SiMe 2CH 2PPh 2]. Addition of unlabelled CO to the specifically labelled benzoyl complex Ni( 13COC 6H 5)[N(SiMe 2CH 2PPh 2) 2] produces Ni(CO) 2[C 6H 5 13 CN(SiMe 2CH 2-PR′ 2)O SiMe 2CH 2PR′ 2] as the only product establishing that no CO insertion into the Ni amide bond is occurring. Kinetic measurements are consistent with an associative pathway for the reductive elimination of the acyl and amide ligands. The X-ray crystal structure of Ni(COC 6H 5)[N(SiMe 2CH 2PPh 2) 2] is reported and shows that the acyl unit is perpendicular to the square-plane of the complex; the NiC bond distance of 1.878 Å is typical of a single bond.

Carbon monoxide metabolism in cell suspensions of Peptostreptococcus productus strain Marburg

Abstract Peptostreptococcus productus strain Marburg is able to grow on CO as the sole energy source and converts this substrate to acetate and CO 2 according to the following equation: 4 CO + 2 H 2 O → 1 acetate − + 1 H + + 2 CO 2 In cell suspensions of the organism the apparent K m value for CO conversion to acetate was about 270 μM (∼ 25% in the gas phase at 140 kPa). Cells incubated in the presence of 14 CO and unlabelled CO 2 incorporated CO specifically into the carboxyl group of acetate like other acetogenic bacteria tested. However, in contrast to experiments performed with other acetogens, 100% of the carboxyl group formed was derived from CO rather than from CO 2 indicating that ‘free’ CO rather than a bound carbonyl formed from CO 2 may be the precursor for the synthesis of the carboxyl group of acetate in this organism.

Determination of substrate specificity of carboxylases by nuclear magnetic resonance

Determination of whether CO 2 or HCO 3 − is the substrate for an enzymatic carboxylation has generally been accomplished by taking advantage of the fact that equilibration of these two compounds requires more than a minute at temperatures below 15°C; thus different kinetics of carboxylation are obtained depending on whether CO 2 or HCO 3 − is used to initiate the reaction. We report a new method using 13C 18O 2 as substrate for determining the CO 2 HCO 3 − specificity of carboxylases. If CO 2 is the substrate, then the 18O content of the 13 C-containing product is the same as that of the 13CO 2 used, whereas if HCO 3 − is the substrate, the 18O content is 2 3 that of the starting material. The method is independent of the detailed kinetics of the CO 2 HCO 3 − interconversion and independent of the presence of contaminating unlabeled CO 2 or HCO 3 −. Isotopic analysis is accomplished by 13C NMR. The method has been used to confirm that HCO 3 − is the substrate for phosphoenolpyruvate carboxylase. Studies of oxygen-18 isotope shifts in phosphorus NMR spectra have permitted confirmation of the observation that label is transferred from HC 18O 3 − into P i during the carboxylation of phosphoenolpyruvate.

Validation of doubly labeled water method using a ruminant.

CO2 production (CDP, ml CO2 . g-1 . h-1) by captive caribou and reindeer (Rangifer tarandus) was measured using the doubly labeled water method (3H2O and H2(18)O) and compared with CO2 expiration rates (VCO2), adjusted for CO2 losses in CH4 and urine, as determined by open-circuit respirometry. CDP calculated from samples of blood or urine from a reindeer in winter was 1-3% higher than the adjusted VCO2. Differences between values derived by the two methods of 5-20% were found in summer trials with caribou. None of these differences were statistically significant (P greater than 0.05). Differences in summer could in part be explained by the net deposition of 3H, 18O, and unlabeled CO2 in antlers and other growing tissues. Total body water volumes calculated from 3H2O dilution were up to 15% higher than those calculated from H2(18)O dilution. The doubly labeled water method appears to be a reasonably accurate method for measuring CDP by caribou and reindeer in winter when growth rates are low, but the method may overestimate CDP by rapidly growing and/or fattening animals.

Anaerobic C 1 Metabolism of the O-Methyl- 14 C-Labeled Substituent of Vanillate

The O-methyl substituents of aromatic compounds constitute a C(1) growth substrate for a number of taxonomically diverse anaerobic acetogens. In this study, strain TH-001, an O-demethylating obligate anaerobe, was chosen to represent this physiological group, and the carbon flow when cells were grown on O-methyl substituents as a C(1) substrate was determined by C radiotracer techniques. O-[methyl-C]vanillate (4-hydroxy-3-methoxy-benzoate) was used as the labeled C(1) substrate. The data showed that for every O-methyl carbon converted to [C]acetate, two were oxidized to CO(2). Quantitation of the carbon recovered in the two products, acetate and CO(2), indicated that acetate was formed in part by the fixation of unlabeled CO(2). The specific activity of C in acetate was 70% of that in the O-methyl substrate, suggesting that only one carbon of acetate was derived from the O-methyl group. Thus, it is postulated that the carboxyl carbon of the product acetate is derived from CO(2) and the methyl carbon is derived from the O-methyl substituent of vanillate. The metabolism of O-[methyl-C]vanillate by strain TH-001 can be described as follows: 3CH(3)OC(7)H(5)O(3) + CO(2) + 4H(2)O --> CH(3)COOH + 2CO(2) + 10H + 10e + 3HOC(7)H(5)O(3).

Solid-state fermentation, lignin degradation and resulting digestibility of wheat straw fermented by selected white-rot fungi

Lignin biodegradation, carbon loss and in vitro dry matter digestibility (IVDMD) have been investigated during the solid state fermentation of wheat straw by eight previously selected strains of white-rot fungi. A mathematical model of the degradation kinetics is presented. [The time period required to reach maximum rates of 14CO2 and unlabeled CO2 release from (14C)-lignin-labelled wheat straw and from whole wheat straw, respectively, was generally short (6–10 days).] High rates of 14C-lignin degradation were achieved by Pycnoporus cinnabarinus (2.9% 14CO2 evolved/day), an unidentified strain Nancon (3.0%/day), Sporotrichum pulverulentum Nov. (3.4%/day), Bjerkandera adusta (2.4%/day), and Dichomitus squalens (2.3%). However, only the latter two strains degraded whole wheat straw slowly and Bjerkandera adusta was not able to degrade more than 23% of the 14C-lignin.

Starch and Sucrose Synthesis in Phaseolus vulgaris as Affected by Light, CO2, and Abscisic Acid

Phaseolus vulgaris L. leaves were subjected to various light, CO(2), and O(2) levels and abscisic acid, then given a 10 minute pulse of (14)CO(2) followed by a 5 minute chase with unlabeled CO(2). After the chase period, very little label remained in the ionic fractions (presumed to be mostly carbon reduction and carbon oxidation cycle intermediates and amino acids) except at low CO(2) partial pressure. Most label was found in the neutral, alcohol soluble fraction (presumed sucrose) or in the insoluble fraction digestable by amyloglucosidase. Sucrose formation was linearly related to assimilation rate (slope = 0.35). Starch formation increased linearly with assimilation rate (slope = 0.56) but did not occur if the assimilation rate was below 4 micromoles per square meter per second. Neither abscisic acid, nor high CO(2) in combination with low O(2) (thought to disrupt control of carbon metabolism) caused significant perturbations of the sucrose/starch formation ratio. These studies indicate that the pathways for starch and sucrose synthesis both are controlled by the rate of net CO(2) assimilation, with sucrose the preferred product at very low assimilation rates.

Sources of Sucrose Translocated from Illuminated Sugar Beet Source Leaves

A search for source leaf sucrose pools that differed in their relation to export was carried out in photosynthesizing leaves of Beta vulgaris L. The time course of depletion of [(14)C]sucrose in a leaf in unlabeled CO(2) following steady state labeling provided evidence for two distinct sucrose pools. After the start of the light period, leaf blade sucrose remained constant although it exchanged between the two pools. Newly synthesized sucrose destined for export passed through one pool more rapidly than through the other. All of the leaf blade sucrose appeared to exchange with export sucrose. Modeling and regression analysis of [(14)C]sucrose data provided a means for estimating the size of the two pools. From 20 to 40% of the sucrose was calculated to be present in the pool that provided the less direct path to export; this was likely vacuolar sucrose. The remainder of the sucrose in the blade was probably in the cytoplasm and veins. Added amounts of leaf blade sucrose, produced in response to elevated CO(2), appeared to be stored mainly in the vacuolar compartment.

CO2 production in animals: analysis of potential errors in the doubly labeled water method.

production are accurate to within &80/c in nine species of mamsumptions associated with this method that, if invalid, mals and reptiles, a bird, and an insect. However, in field could cause errors in calculated metabolic rates. These studies, errors can be much larger under certain circumstances. assumptions, as listed by Lifson and McClintock (16) are: Isotopic fraction of labeled water can cause large errors in I) body water volume remains constant during the measanimals whose evaporative water loss comprises a major prourement period, 2) rates of water flux and CO2 production portion of total water efflux. Input of CO2 across lungs and skin are constant through time, 3) the isotopes label only the caused errors exceeding +80’S in kangaroo rats exposed to air Hz0 and CO* in the body, 4) the isotopes leave the body containing 3.4% unlabeled CO?. Analytical errors of HYo in isot,ope concentrations can cause calculated rates of COY proonly as Hz0 and COZ, 5) the specific activities of the

Heterogeneous response of hepatic mixed function oxidases to chronic phenobarbital administration

Effects of chronic phenobarbital (PB) administration (75mg/kg, p.o. for 59 days) on certain hepatic mixed function oxidases (HMFO) were investigated in male Sprague-Dawley rats to determine whether maximal induction is maintained throughout this period or whether further alterations occur in the inductive effect. As demonstrated by measurements of 14CO 2 excretion rates in breath after a singl i.p. injection of [dimethylamine- 14C]aminopyrine ([ 14C]aminopyrine), PB in the above dose for 7 days accelerated hepatic aminopyrine demethylation 92 per cent above control values. However, after 59 days of PB administration, 14CO 2 excretion was only 5 per cent above control values; thus, initial PB-induced enhancement of aminopyrine N-demethylation declined with time. Phenobarbital administration did not affect total (unlabeled) CO 2 output, indicating that decreased 14CO 2 excretion did not result from potential PB-induced reduction of CO 2 output. In contrast, total cytochrome P450 content and aniline hydroxylase activity remained elevated throughout the 59 days of PB treatment. Ethylmorphine N-demethylase activity and aminopyrine N-demethylase activity (using a 3.0 mM substrate concentration) remained elevated until 28 days of PB administration, after which time these activities decreased significantly (P < 0.05), finally reaching 71 and 83 per cent respectively, of peak activity. Kinetic analyses of aminopyrine N-demethylation by hepatic microsomes revealed the K m values were unchanged from 4 to 59 days of PB administration, but that V max increased to a maximum of 15.5 on day 28, after which it decreased to 6.60 on day 59. This pattern of change for V max of aminopyrine N-demethylase was similar to that for enzyme activity obtained using a constant 3.0 mM concentration of aminopyrine. Although aminopyrine N-demethylase activity (using a 3.0 mM substrate concentration) peaked after 2 days of PB treatment, 7 days of PB treatment reduced this activity to 14 per cent below peak values. After 59 days of PB treatment, aminopyrine N-demethylase activity (at a 0.3 mM substrate concentration) declined to 61 per cent of the peak value attained on day 2. Another chronic PB study was performed and different time points selected to determine the reproducibility of these differential alterations of MFO activity. Results in the second study were similar to those obtained in the first study. Appreciable loss with time of the inductive effects produced by PB on aminopyrine metabolism both in vitro and in vivo suggests a heterogeneous response of hepatic MFO to PB; metabolism of aniline remained at peak inductive values throughout the 59 days of PB administration, whereas ethylmorphine metabolism declined only slightly.

Glutamine-dependent carbamyl phosphate synthetase: catalysis and regulation

Glutamine-dependent carbamyl phosphate synthetase: catalysis and regulation

Seasonal transfers of assimilated 14C in grassland: Plant production and turnover, soil and plant respiration

Six areas of native grassland were labelled with 14C during a growing season. Transfers from the foliage to the roots and root respiration were measured. Plant production and turnover rates were determined by sampling the labelled material at different periods following exposure to 14CO 2. Above to beneath ground plant production ratios ranged between 1.1 and 1.9 with maximal translocation to the roots occurring during the drier summer months. The distribution of the photosynthates in the roots at different depths changed with time and soil moisture content. The upper part of the soil (0–10 cm) contained 49–77% of the labelled C found beneath the soil surface. Measurement of transfers with time of the above ground labelled C from living to dead plant and litter categories gave an insight into foliage dynamics and made it possible to estimate the seasonal shoot production at 130g Cm −2 (1300kg ha −1). Root growth represented 100g Cm −2 (1000 kg ha −1). Calculations of root and soil respiration were based on the CO 2 profiles in the soil. The fluxes of labelled and unlabelled CO 2 at the soil surface were estimated using the diffusion equation method. Respiration by roots and closely associated soil organisms accounted for 12 per cent of the net assimilation of CO 2 by the plants. This proportion was constant throughout the season and represented 19 per cent of the total CO 2 evolved at the soil surface.

Carbon dioxide fixation by Veillonella parvula M 4 and its relation to propionic acid formation.

The effect of carbon dioxide on propionate formation during lactate and pyruvate metabolism by intact cells and cell-free preparations of Veillonella parvula M4 was examined. While the overall rate of lactate-U-14C degradation by intact cells was not altered, concentrations of unlabeled CO2 (to 50%) stimulated the evolution of 14CO2, and increased the absolute quantity of the propionate formed, while decreasing its specific activity. Acetate production from the labeled lactate, however, was not affected. Furthermore, propionate formation was not dependent on the presence of CO2 in the gas phase. The principle product of pyruvate-3-14C metabolism by extracts of the organism was acetate, although minor amounts of labeled propionate were also formed. Small quantities of oxaloacetate were detected during this metabolism by direct enzymatic assay and by separation of its hydrazone by thin-layer chromatography. Dialyzed crude extracts of the organism, treated with protamine sulfate and charcoal, were shown to catalyze the formation of oxaloacetate from pyruvate and CO2 in the presence of ATP when assayed both enzymatically with malic dehydrogenase and NADH, and with pyruvate-3-14C. This CO2-fixing activity could be attributed to pyruvate carboxylase, since the activity was Mg2+-dependent, was inhibited by avidin and aspartate, and was stimulated by biotin and CoA. Furthermore, the same extract preparation was capable of converting pyruvate-3-14C and CO2 to malate in the presence of NADH and NADPH indicating the presence of malic enzyme in the organism; this activity was confirmed by spectrophotometric means. As is the case with V. alcalescens, dialyzed extracts of V. parvula M4 catalyzed the exchange reaction between the carboxyl group of pyruvate and CO2; this exchange activity was stimulated by ATP.

Studies on the Mechanism of Glycerate 3-Phosphate Synthesis in Tomato and Maize Leaves

The net carbon incorporation in maize (Zea mays) and tomato (Lycopersicum esculentum) leaves was mainly the result of the carboxylation of ribulose 1,5-diphosphate. In both of these organisms synthesis of glycerate 3-phosphate was studied during short chase experiments (2 or 3 seconds in (14)CO(2) then 8 to 27 seconds in unlabeled CO(2)). Changes in the radioactivity in the individual carbon atoms of glycerate 3-phosphate, malate, and aspartate are consistent with the formation, in both leaves, of 2 molecules of glycerate 3-phosphate for each CO(2) molecule incorporated. The CO(2), before reacting with ribulose 1,5-diphosphate, is first incorporated in an intracellular CO(2) pool which has a different composition according to the species. This pool is constituted in tomato by volatile compounds (50 nanomoles per gram of fresh weight) more or less in equilibrium with atmospheric CO(2). In maize the pool consists of carbon atoms 4 of malate and aspartate (for at least 80% of the pool) and volatile compounds which correspond, in all, to 540 nanomoles per gram of fresh weight where atmospheric CO(2) enters through an irreversible reaction.