Carbon isotope fractionation during photorespiration and carboxylation in Senecio

The magnitude of fractionation during photorespiration and the effect on net photosynthetic (13)C discrimination (Delta) were investigated for three Senecio species, S. squalidus, S. cineraria, and S. greyii. We determined the contributions of different processes during photosynthesis to Delta by comparing observations (Delta(obs)) with discrimination predicted from gas-exchange measurements (Delta(pred)). Photorespiration rates were manipulated by altering the O(2) partial pressure (pO(2)) in the air surrounding the leaves. Contributions from (13)C-depleted photorespiratory CO(2) were largest at high pO(2). The parameters for photorespiratory fractionation (f), net fractionation during carboxylation by Rubisco and phosphoenolpyruvate carboxylase (b), and mesophyll conductance (g(i)) were determined simultaneously for all measurements. Instead of using Delta(obs) data to obtain g(i) and f successively, which requires that b is known, we treated b, f, and g(i) as unknowns. We propose this as an alternative approach to analyze measurements under field conditions when b and g(i) are not known or cannot be determined in separate experiments. Good agreement between modeled and observed Delta was achieved with f = 11.6 per thousand +/- 1.5 per thousand, b = 26.0 per thousand +/- 0.3 per thousand, and g(i) of 0.27 +/- 0.01, 0.25 +/- 0.01, and 0.22 +/- 0.01 mol m(-2) s(-1) for S. squalidus, S. cineraria, and S. greyii, respectively. We estimate that photorespiratory fractionation decreases Delta by about 1.2 per thousand on average under field conditions. In addition, diurnal changes in Delta are likely to reflect variations in photorespiration even at the canopy level. Our results emphasize that the effects of photorespiration must be taken into account when partitioning net CO(2) exchange of ecosystems into gross fluxes of photosynthesis and respiration.     

13C isotope fractionation during rhizosphere respiration of C3 and C4 plants

Stable carbon isotopes are used extensively to partition total soil CO₂ efflux into root-derived rhizosphere respiration or autotrophic respiration and soil-derived heterotrophic respiration. However, it remains unclear whether CO₂ from rhizosphere respiration has the same δ¹³C value as root biomass. Here we investigated the magnitude of ¹³C isotope fractionation during rhizosphere respiration relative to root biomass in six plant species. Plants were grown in a carbon-free sand-perlite medium inoculated with microorganisms from a farm soil for 62 days inside a greenhouse. We measured the δ¹³C value of rhizosphere respiration using a closed-circulation 48-hour CO₂ trapping method during 40~42 and 60~62 days after sowing. We found a consistent depletion in ¹³C (0.9~1.7‰) of CO₂ from rhizosphere respiration relative to root biomass in three C₃ species (Glycine max L. Merr., Helianthus annuus L. and Triticum aestivum L.), but a relatively large depletion in ¹³C (3.7~7.0‰) in three C₄ species (Amaranthus tricolor L., Sorghum bicolor (L.) Moench and Zea mays L. ssp. mays). Overall, our results indicate that CO₂ from rhizosphere respiration is more ¹³C-depleted than root biomass. Therefore, accounting for this ¹³C fractionation is required for accurately partitioning total soil CO₂ efflux into root-derived and soil-derived components using natural abundance stable carbon isotope methods.     

Carbon isotope fractionation in plants

Plants with the C₃, C₄, and crassulacean acid metabolism (CAM) photosynthetic pathways show characteristically different discriminations against ¹³C during photosynthesis. For each photosynthetic type, no more than slight variations are observed within or among species. CAM plants show large variations in isotope fractionation with temperature, but other plants do not. Different plant organs, subcellular fractions and metabolises can show widely varying isotopic compositions. The isotopic composition of respired carbon is often different from that of plant carbon, but it is not currently possible to describe this effect in detail. The principal components which will affect the overall isotope discrimination during photosynthesis are diffusion of CO₂, interconversion of CO₂ and HCO^?₃, incorporation of CO₂ by phosphoenolpyruvate carboxylase or ribulose bisphosphate carboxylase, and respiration. Theisotope fractionations associated with these processes are summarized. Mathematical models are presented which permit prediction of the overall isotope discrimination in terms of these components. These models also permit a correlation of isotope fractionations with internal CO₂ concentrations. Analysis of existing data in terms of these models reveals that CO₂ incorporation in C₃ plants is limited principally by ribulose bisphosphate carboxylase, but CO₂ diffusion also contributes. In C₄ plants, carbon fixation is principally limited by the rate of CO₂ diffusion into the leaf. There is probably a small fractionation in C₄ plants due to ribulose bisphosphate carboxylase.     

Relationship between transpiration and respiration in plants during the dark period

Energetic aspects of the relation between transpiration and respiration during the dark period were evaluated. One-year old seedlings of three trees, one bush and one annual plant were grown in controlled conditions. Experiments were performed under uniform environment during the day and two regimes of air relative humidity (RH) during the night, low (50 - 65 %) and high (95 %). For all investigated plant species the dark transpiration rate (E), the free energy of respiratory substrate, the entropy production and the free energy balance (FEB) of the dark respiration were higher at low than at high RH. E was linearly related to the FEBr ² ranged between 0.63 and 0.90)     

Calcium isotope fractionation in alpine plants

In order to develop Ca isotopes as a tracer for biogeochemical Ca cycling in terrestrial environments and for Ca utilisation in plants, stable calcium isotope ratios were measured in various species of alpine plants, including woody species, grasses and herbs. Analysis of plant parts (root, stem, leaf and flower samples) provided information on Ca isotope fractionation within plants and seasonal sampling of leaves revealed temporal variation in leaf Ca isotopic composition. There was significant Ca isotope fractionation between soil and root tissue $\Updelta^{44/42}\hbox{Ca}_{\rm root-soil} \approx -0.40\,\permille$ in all investigated species, whereas Ca isotope fractionation between roots and leaves was species dependent. Samples of leaf tissue collected throughout the growing season also highlighted species differences: Ca isotope ratios increased with leaf age in woody species but remained constant in herbs and grasses. The Ca isotope fractionation between roots and soils can be explained by a preferential binding of light Ca isotopes to root adsorption sites. The observed differences in whole plant Ca isotopic compositions both within and between species may be attributed to several potential factors including root cation exchange capacity, the presence of a woody stem, the presence of Ca oxalate, and the levels of mycorrhizal infection. Thus, the impact of plants on the Ca biogeochemical cycle in soils, and ultimately the Ca isotope signature of the weathering flux from terrestrial environments, will depend on the species present and the stage of vegetation succession.     

Carbon nutrition of seaweeds: Photosynthesis,photorespiration and respiration

Techniques are described for measuring gas exchange in seaweeds held in moist air (air suspension). In the species we have examined, oxygen has little or no effect on photosynthesis except at very low (50 μ1·1^?1) CO₂ concentration. Photorespiration could not be detected unless the seaweeds were treated simultaneously with high O₂ and low CO₂ or with the carbonic anhydrase inhibitor, diamox. However, sporulating and meristematic tissues exhibit oxygen-insensitive light respiration (CO₂production in light not associated with photorespiratory metabolism). Elevated pH in the surface water of seaweeds also caused light respiration. Oxygen-sensitive wound respiration was observed that could easily be mistaken for photorespiration. C₄ photosynthesis could not be detected. On the basis of several experimental approaches it was concluded that these seaweeds normally absorb bicarbonate rather than CO₂ from sea water. High CO₂ concentrations are required in gas streams aerating seaweed cultures in air or water suspension to maintain the bicarbonate concentration at levels normally found in sea water and to support normal levels of photosynthesis.     

Specific features of photorespiration in photosynthetically active organs of C3 plants

Photorespiration of photosynthetically active organs of C₃ plants (leaf, ear, stem, and leaf sheath) and C₄ plants (leaf, tassel, stem, leaf sheath, ear husk) grown under greenhouse and field conditions was studied. Photorespiration was measured using a PTM-48A high-technology monitor of photosynthesis (Bioinstruments S.R.L., Moldova). It is shown that photorespiration (CO₂ ejection after light turning off — apparent photorespiration) in C₃ plants is characteristic only for their leaves. In other photosynthesizing organs, photorespiration was absent, like in the photosynthesizing organs of C₄ plants. The absence of such after-light CO₂ outburst was observed for 31 genotypes: 18 cereal species belonging to four species (Triticum aestivum L., T. durum Desf., Secale cereale L., and Triticale); 6 grain legumes belonging to 2 species (Pisum sativum L. and Glycine max L.); 7 species of wild and rarely cultivated genotypes (T. boeoticum Boiss., T. dicoccoides Koern., T. dicoccum Schuebl., T. spelta L., T. compactum Host., T. monococcum L., and T. sphaerococcum Persiv.), and 2 genotypes of C₄ plants (Zea mays L. and Sorgum vulgaris L.). In all tested photosynthetically active genotypes, except of the C₃ plant leaves, apparent photorespiration was absent, but rather active glycolate cycle operated. The activity of this cycle was determined from the activity of the key enzyme of this cycle — glycolate oxidase. It was supposed that C₃ plants have two mechanisms of CO₂ assimilation: the first one — the mechanism of C₃ type localized in the leaves and the second one localized in other photosynthesizing organs, similar or with some elements of C₄ mechanism of CO₂ assimilation, limiting after-light CO₂ ejection during the metabolism of glycolate.     

Mg and Ca isotope fractionation during CaCO3 biomineralisation

The natural variation of Mg and Ca stable isotopes of carbonates has been determined in carbonate skeletons of perforate foraminifera and reef coral together with Mg/Ca ratios to assess the influence of biomineralisation processes. The results for coral aragonite suggest its formation, in terms of stable isotope behaviour, approximates to inorganic precipitation from a seawater reservoir. In contrast, results for foraminifera calcite suggest a marked biological control on Mg isotope ratios presumably related to its low Mg content compared with seawater. The bearing of these observations on the use of Mg and Ca isotopes as proxies in paleoceanography is considered.     

Light-enhanced dark respiration in leaves and mesophyll protoplasts of pea in relation to photorespiration,respiration and some metabolites content

The respiration rate of leaves and mesophyll protoplasts of pea (Pisum sativum L.), from plants which were previously kept in darkness for 24 h was doubled following a period of photosynthesis at ambient level of O₂ (21 %), whereas the low level of O₂ (1 % and 4 % for leaves and protoplasts, respectively) reduced this light-enhanced dark respiration (LEDR) to the rate as noted before the illumination. Similarly to respiration rate, the oxygen at used concentrations had no effect on the ATP/ADP ratio in the dark-treated leaves. However, the ATP/ADP ratio in leaves photosynthesizing at 21 % O₂ was higher (up to 40 %, dependence on CO₂ concentration in the range 40–1600 1 dm⁻³) than in those photosynthesizing at 1 % O₂ or darkened at air (21 % O₂). Also, at 1 % O₂ the accumulation of malate was suppressed (by about 40 %), to a value noted for leaves darkened at 21 % O₂. The dark-treatment of leaves reduced the ability of isolated mitochondria to oxidize glycine (by about twofold) and succinate, but not malate. Mitochondria from both the light- and dark-treated leaves did not differ in qualitative composition of free amino acids, however, there were significant quantitative differences especially with respect to aspartate, alanine, glutamate and major intermediates of the photorespiratory pathway (glycine, serine). Our results suggest that accumulation of photorespiratory and respiratory metabolites in pea leaves during photosynthesis at 1 % O₂ is reduced, hence the suppression of postillumination respiration rate.     

Short-term measurement of carbon isotope fractionation in plants

Combustion-based studies of the carbon-13 content of plants give only an integrated, long-term value for the isotope fractionation associated with photosynthesis. A method is described here which permits determination of this isotope fractionation in 2 to 3 hours. To accomplish this, the plant is enclosed in a glass chamber, and the quantity and isotopic content of the CO₂ remaining in the atmosphere are monitored during photosynthesis. Isotope fractionation studies by this method give results consistent with what is expected from combustion studies of C₃, C₄, and Crassulacean acid metabolism plants. This method will make possible a variety of new studies of environmental and species effects in carbon isotope fractionation.     

Maintenance and growth components of dark respiration rate in leaves of C3 and C4 plants as affected by leaf temperature

The rates of maintenance and growth components of leaf dark respiration of a C₃ plant (Phaseolus vulgaris L.) and C₄ plant (Zea mays L.) as affected by temperature were studied using the McCree concept. Respiration rates were measured by means of infrared gas analysis in a closed gas exchange system. In both C₃ and C₄ species R_D and R_m increased with temperature in the temperature range (15–62 °C) studied. R_G depended on temperature with an optimum near the temperature optimum of gross photosynthetic rate, P_g. Significant correlation between R_D and R_M and between R_G and P_G was found.     

C-isotope composition of CO2 respired by shoots and roots: fractionation during dark respiration?

The CO₂ respired by leaves is ¹³C-enriched relative to leaf biomass and putative respiratory substrates (Ghashghaie et al., Phytochemistry Reviews 2, 145–161, 2003), but how this relates to the ¹³C content of root, or whole plant respiratory CO₂ is unknown. The C isotope composition of respiratory CO₂ (δ_R) from shoots and roots of sunflower (Helianthus annuus L.), alfalfa (Medicago sativa L.), and perennial ryegrass (Lolium perenne L.) growing in a range of conditions was analysed. In all instances plants were grown in controlled environments with CO₂ of constant concentration and δ¹³C. Respiration of roots and shoots of individual plants was measured with an open CO₂ exchange system interfaced with a mass spectrometer. Respiratory CO₂ from shoots was always ¹³C-enriched relative to that of roots. Conversely, shoot biomass was always ¹³C-depleted relative to root biomass. The δ-difference between shoot and root respiratory CO₂ was variable, and negatively correlated with the δ-difference between shoot and root biomass (r² = 0.52, P = 0.023), suggesting isotope effects during biosynthesis. ¹³C discrimination in respiration (R) of shoots, roots and whole plants (e_Shoot, e_Root, e_Plant) was assessed as e = (δ_Substrate − δ_R)/(1 + δ_R/1000), where root and shoot substrate is defined as imported C, and plant substrate is total photosynthate. Estimates were obtained from C isotope balances of shoots, roots and whole plants of sunflower and alfalfa using growth and respiration data collected at intervals of 1 to 2 weeks. e_plant and e_Shoot differed significantly from zero. e_plant ranged between −0.4 and −0.9‰, whereas e_Shoot was much greater (−0.6 to −1.9‰). e_Root was not significantly different from zero. The present results help to resolve the apparent conflict between leaf- and ecosystem-level ¹³C discrimination in respiration.     

Carbon and hydrogen stable isotope fractionation during aerobic bacterial degradation of aromatic hydrocarbons

13C/(12)C and D/H stable isotope fractionation during aerobic degradation was determined for Pseudomonas putida strain mt-2, Pseudomonas putida strain F1, Ralstonia pickettii strain PKO1, and Pseudomonas putida strain NCIB 9816 grown with toluene, xylenes, and naphthalene. Different types of initial reactions used by the respective bacterial strains could be linked with certain extents of stable isotope fractionation during substrate degradation.     

Physical and chemical basis of carbon isotope fractionation in plants

Naturally-occurring variations in the abundances of the stable isotopes of carbon and other elements can be used to understand the dynamics of natural processes in chemistry, biochemistry, biology, medicine, ecology and other fields. The use of carbon-13 isotopic abundances as an indicator of photosynthetic function in plants has become common. The purpose of this article is to describe the physical and chemical processes that contribute to the abundances of carbon-13 in plant materials, and to provide a framework for understanding how those processes control the isotopic contents of natural materials.     

13C/12C isotope fractionation of aromatic hydrocarbons during microbial degradation

The influence of microbial degradation on the ¹³C/¹²C isotope composition of aromatic hydrocarbons is presented using toluene as a model compound. Four different toluene-degrading bacterial strains grown in batch culture with oxygen, nitrate, ferric iron or sulphate as electron acceptors were studied as representatives of different environmental redox conditions potentially prevailing in contaminated aquifers. The biological degradation induced isotope shifts in the residual, non-degraded toluene fraction and the kinetic isotope fractionation factors αC for toluene degradation by Pseudomonas putida (1.0026 ± 0.00017), Thauera aromatica (1.0017 ± 0.00015), Geobacter metallireducens (1.0018 ± 0.00029) and the sulphate-reducing strain TRM1 (1.0017 ± 0.00016) were in the same range for all four species, although they use at least two different degradation pathways. A similar ¹³C/¹²C isotope fractionation factor (αC = 1.0015 ± 0.00015) was observed in situ in a non-sterile soil column in which toluene was degraded under sulphate-reducing conditions. No carbon isotope shifts resulting from soil–hydrocarbon interactions were observed in a non-degrading soil column control with aquifer material under the same conditions. The results imply that microbial degradation of toluene can produce a ¹³C/¹²C isotope fractionation in the residual hydrocarbon fraction under different environmental conditions.     

Interaction between photorespiration and respiration in transgenic potato plants with antisense reduction in glycine decarboxylase

Potato (Solanum tuberosum L. cv. Désirée) plants with an antisense reduction in the P-protein of the glycine decarboxylase complex (GDC) were used to study the interaction between respiration and photorespiration. Mitochondria isolated from transgenic plants had a decreased capacity for glycine oxidation and glycine accumulated in the leaves. Malate consumption increased in leaves of GDC deficient plants and the capacity for malate and NADH oxidation increased in isolated mitochondria. A lower level of alternative oxidase protein and decreased partitioning of electrons to the alternative pathway was found in these plants. The adenylate status was altered in protoplasts from transgenic plants, most notably the chloroplastic ATP/ADP ratio increased. The lower capacity for photorespiration in leaves of GDC deficient plants was compensated for by increased respiratory decarboxylations in the light. This is interpreted as a decreased light suppression of the tricarboxylic acid cycle in GDC deficient plants in comparison to wild-type plants. The results support the view that respiratory decarboxylations in the light are restricted at the level of the pyruvate dehydrogenase complex and/or isocitrate dehydrogenase and that this effect is likely to be mediated by mitochondrial photorespiratory products.     

Effect of phosphinothricin (glufosinate) on photosynthesis and photorespiration of C3 and C4 plants

Phosphinothricin (glufosinate), an irreversible inhibitor of glutamine synthetase, causes an inhibition of photosynthesis in C₃ (Sinapis alba) and C₄ (Zea mays) plants under atmospheric conditions (400 ppm CO₂, 21% O₂). This photosynthesis inhibition is proceeding slower in C₄ leaves. Under non-photorespiratory conditions (1000 ppm CO₂, 2% O₂) there is no inhibition of photosynthesis. The inhibition of glutamine synthetase by phosphinothricin results in an accumulation of NH₄ ⁺. The NH₄ ⁺-accumulation is lower in C₄ plants than in C₃ plants. The inhibition of glutamine synthetase through phosphinothricin in mustard leaves results in a decrease in glutamine, glutamate, aspartate, asparagine, serine, and glycine. In contrast to this, a considerable increase in leucine and valine following phosphinothricin treatment is measured. With the addition of either glutamine, glutamate, aspartate, glycine or serine, photosynthesis inhibition by phosphinothricin can be reduced, although the NH₄ ⁺-accumulation is greatly increased. This indicates that NH₄ ⁺-accumulation cannot be the primary cause for photosynthesis inhibition by phosphinothricin. The investigations demonstrate the inhibition of transmination of glyoxylate to glycine in photorespiration through the total lack of amino donors. This could result in a glyoxylate accumulation inhibiting ribulose-1,5-bisphosphate-carboxylase and consequently CO₂-fixation.Abbreviations GOGAT glutamine-2-oxoglutarate-amidotransferase - GS glutamine synthetase - PPT phosphinothricin - MSO methionine sulfoximine - RuBP ribulose-1,5-bisphosphate     

Effects of light on respiration and oxygen isotope fractionation in soybean cotyledons

Light effects on electron flow through the cyanide-resistant respiratory pathway, oxygen isotope fractionation and total respiration were studied in soybean (Glycine max L.) cotyledons. During the first 12 h of illumination there was an increase in both electron partitioning through the alternative pathway and oxygen isotope fractionation by the alternative oxidase. The latter probably indicates a change in the properties of the alternative oxidase. There was no engagement of the alternative oxidase in darkness and its fractionation was 27‰. In green cotyledons 60% of the respiration flux was through the alternative pathway and the alternative oxidase fractionation was 32‰. Exposing previously illuminated tissue to continuous darkness induced a decrease in the electron partitioning through the alternative pathway. However, this decrease was not directly linked with the low cellular sugar concentration resulting from the lack of light because 5 min of light every 12 h was sufficient to keep the alternative pathway engaged to the same extent as plants grown under control conditions.     

Carbon isotope fractionation during acetoclastic methanogenesis by Methanosaeta concilii in culture and a lake sediment

The isotope enrichment factors (epsilon) in Methanosaeta concilii and in a lake sediment, where acetate was consumed only by Methanosaeta spp., were clearly less negative than the epsilon usually observed for Methanosarcina spp. The fraction of methane produced from acetate in the sediment, as determined by using stable isotope signatures, was 10 to 15% lower when the appropriate epsilon of Methanosaeta spp. was used.