Carbon isotope fractionation by ribulose-1,5-bisophosphate carboxylase from various organisms

Carbon isotope fractionation by structurally and catalytically distinct ribulose-1,5-bisphosphate carboxylases from one eucaryotic and four procaryotic organisms has been measured under nitrogen. The average fractionation for 40 experiments was −34.1 ‰ with respect to the δ¹³C of the dissolved CO₂ used, although average fractionations for each enzyme varied slightly: spinach carboxylase, −36.5 ‰; Hydrogenomonas eutropha, −38.7 ‰; Agmenellum quadruplicatum, −32.2 ‰; Rhodospirillum rubrum, −32.1 ‰; Rhodopseudomonas sphaeroides peak I carboxylase, −31.4 ‰; and R. sphaeroides peak II carboxylase, −28.3 ‰. The carbon isotope fractionation value was largely independent of method of enzyme preparation, purity, or reaction temperature, but in the case of spinach ribulose-1,5-bisphosphate carboxylase fractionation, changing the metal cofactor used for enzyme activation had a distinct effect on the fractionation value. The fractionation value of −36.5 ‰ with Mg²⁺ as activator shifted to −29.9 ‰ with Ni²⁺ as activator and to −41.7 ‰ with Mn²⁺ as activator. These dramatic metal effects on carbon isotope fractionation may be useful in examining the catalytic site of the enzyme.     

Carbon isotope ratios in crassulacean Acid metabolism plants: seasonal patterns from plants in natural stands

A year round study of photosynthesis and carbon isotope fractionation was conducted with plants of Opuntia phaeacantha Engelm. and Yucca baccata Torr. occurring in natural stands at elevations of 525, 970, 1450 and 1900 m. Plant water potentials and the daytime pattern of ¹⁴CO₂ photosynthesis were similar for all cacti along the elevational gradient, despite significant differences in temperature regime and soil water status. Carbon isotope ratios of total tissue and soluble extract fractions were relatively constant throughtout the entire year. Additionally, the σ¹³C values were similar in all plants of the same species along the elevational gradient, i.e. −12.5 ± 0.86 ‰ for O. phaeacantha and −15.7 ± 0.95 ‰ for Y. baccata. The results of this study indicate Crassulacean acid metabolism predominates as the major carbon pathway of these plants, which do not facultatively utilize the reductive pentose phosphate cycle of photosynthesis as the primary carboxylation reaction.     

Carbon Isotope Fractionation of 11 Acetogenic Strains Grown on H2 and CO2

Acetogenic bacteria are able to grow autotrophically on hydrogen and carbon dioxide by using the acetyl coenzyme A (acetyl-CoA) pathway. Acetate is the end product of this reaction. In contrast to the fermentative route of acetate production, which shows almost no fractionation of carbon isotopes, the acetyl-CoA pathway has been reported to exhibit a preference for light carbon. In Acetobacterium woodii the isotope fractionation factor (ε) for ¹³C and ¹²C has previously been reported to be ε = −58.6‰. To investigate whether such a strong fractionation is a general feature of acetogenic bacteria, we measured the stable carbon isotope fractionation factor of 10 acetogenic strains grown on H₂ and CO₂. The average fractionation factor was ε_TIC = −57.2‰ for utilization of total inorganic carbon and ε_acetate = −54.6‰ for the production of acetate. The strongest fractionation was found for Sporomusa sphaeroides (ε_TIC = −68.3‰), the lowest fractionation for Morella thermoacetica (ε_TIC = −38.2‰). To investigate the reproducibility of our measurements, we determined the fractionation factor of 21 biological replicates of Thermoanaerobacter kivui. In general, our study confirmed the strong fractionation of stable carbon during chemolithotrophic acetate formation in acetogenic bacteria. However, the specific characteristics of the bacterial strain, as well as the cultural conditions, may have a moderate influence on the overall fractionation.     

Isotope Fractionation in Photosynthetic Bacteria during Carbon Dioxide Assimilation

The δ _PDB¹³C values have been determined for the cellular constituents and metabolic intermediates of autotrophically grown Chromatium vinosum. The isotopic composition of the HCO₃^- in the medium and the carbon isotopic composition of the bacterial cells change with the growth of the culture. The δ _PDB¹³C value of the HCO₃^- in the media changes from an initial value of −6.6‰ to +8.1‰ after 10 days of bacterial growth and the δ _PDB¹³C value of the bacterial cells change from −37.5‰ to −29.2‰ in the same period. The amount of carbon isotope fractionation during the synthesis of hexoses by the photoassimilation of CO₂ has a range of −15.5‰ at time zero to −22.0‰ after 10 days. This range of fractionation compares to the range of carbon isotope fractionation for the synthesis of sugars from CO₂ by ribulose 1,5-diphosphate carboxylase and the Calvin cycle.     

Stable Carbon Isotope Fractionation by Methylotrophic Methanogenic Archaea

In natural environments methane is usually produced by aceticlastic and hydrogenotrophic methanogenic archaea. However, some methanogens can use C₁ compounds such as methanol as the substrate. To determine the contributions of individual substrates to methane production, the stable-isotope values of the substrates and the released methane are often used. Additional information can be obtained by using selective inhibitors (e.g., methyl fluoride, a selective inhibitor of acetoclastic methanogenesis). We studied stable carbon isotope fractionation during the conversion of methanol to methane in Methanosarcina acetivorans, Methanosarcina barkeri, and Methanolobus zinderi and generally found large fractionation factors (−83‰ to −72‰). We further tested whether methyl fluoride impairs methylotrophic methanogenesis. Our experiments showed that even though a slight inhibition occurred, the carbon isotope fractionation was not affected. Therefore, the production of isotopically light methane observed in the presence of methyl fluoride may be due to the strong fractionation by methylotrophic methanogens and not only by hydrogenotrophic methanogens as previously assumed.     

Effect of Substrate Concentration on Carbon Isotope Fractionation during Acetoclastic Methanogenesis by Methanosarcina barkeri and M. acetivorans and in Rice Field Soil

Methanosarcina is the only acetate-consuming genus of methanogenic archaea other than Methanosaeta and thus is important in methanogenic environments for the formation of the greenhouse gases methane and carbon dioxide. However, little is known about isotopic discrimination during acetoclastic CH₄ production. Therefore, we studied two species of the Methanosarcinaceae family, Methanosarcina barkeri and Methanosarcina acetivorans, and a methanogenic rice field soil amended with acetate. The values of the isotope enrichment factor (ɛ) associated with consumption of total acetate (ɛ_ac), consumption of acetate-methyl (ɛ_ac-methyl) and production of CH₄ (ɛ_CH4) were an ɛ_ac of −30.5‰, an ɛ_ac-methyl of −25.6‰, and an ɛ_CH4 of −27.4‰ for M. barkeri and an ɛ_ac of −35.3‰, an ɛ_ac-methyl of −24.8‰, and an ɛ_CH4 of −23.8‰ for M. acetivorans. Terminal restriction fragment length polymorphism of archaeal 16S rRNA genes indicated that acetoclastic methanogenic populations in rice field soil were dominated by Methanosarcina spp. Isotope fractionation determined during acetoclastic methanogenesis in rice field soil resulted in an ɛ_ac of −18.7‰, an ɛ_ac-methyl of −16.9‰, and an ɛ_CH4 of −20.8‰. However, in rice field soil as well as in the pure cultures, values of ɛ_ac and ɛ_ac-methyl decreased as acetate concentrations decreased, eventually approaching zero. Thus, isotope fractionation of acetate carbon was apparently affected by substrate concentration. The ɛ values determined in pure cultures were consistent with those in rice field soil if the concentration of acetate was taken into account.Methane (CH₄) is the most abundant reduced gas in the earth''s atmosphere and is an important greenhouse gas with a high global-warming potential (7). It is presently a matter of discussion whether the contribution of CH₄ to the greenhouse effect will increase in the future (3, 23). This has made it necessary and more urgent to understand natural processes that lead to the production of CH₄.Methanogenesis, the microbial formation of CH₄, is the final step in the degradation of organic matter in anoxic environments like natural wetlands, lake sediments, and flooded rice fields. The most important precursors for the production of CH₄ are acetate (equation 1) and CO₂ (equation 2) with the following reactions (8): (1) (2)Acetate is the most important substrate since it contributes more than 67% to microbial methanogenesis during anoxic degradation of polysaccharides. In methanogenic environments only two genera of archaea, Methanosaeta and Methanosarcina, are capable of using acetate (2). While Methanosaeta can be considered a specialist that uses only acetate, Methanosarcina can use a wide range of substrates besides acetate, for example, H₂/CO₂, methanol, methylamines, and methylated sulfides. Among methanogens, Methanosarcinaceae also display the largest environmental distribution. They can be found in freshwater sediments and soil, marine habitats, landfills, and animal gastrointestinal tracts (46).Additionally, differences between Methanosarcina and Methanosaeta were found for isotope fractionation of stable carbon. The fractionation factor (α) or, equivalently, the enrichment factor (ɛ) during acetoclastic methanogenesis in Methanosarcina barkeri strains typically ranges from an α of 1.021 to 1.027 or an ɛ of −27‰ to −21‰ (14, 27, 48), whereas isotope fractionation in Methanosaeta spp. is weaker, i.e., an α of 1.007 (ɛ = −7‰) for Methanosaeta thermophila (43) and an α of 1.010 (ɛ = −10‰) for Methanosaeta concilii (34). It is suggested that the two archaeal genera differ in isotope fractionation due to differences in their biochemical activation of acetate to acetyl-coenzyme A (acetyl-CoA) (34). However, isotopic data for acetoclastic methanogens are rare. For instance, all data for Methanosarcina refer to only one species, namely M. barkeri.Hence, in this study we investigated whether differences in carbon isotope fractionation within the genus Methanosarcina occur. Therefore, we determined isotope ratios of stable carbon in cultures of the acetoclastic species M. barkeri and Methanosarcina acetivorans. Second, we were interested if these data, obtained from pure cultures, could also be applied to understand natural environments. For that reason, we determined isotope fractionation during acetoclastic methanogenesis in the model system rice field soil. Furthermore, we discuss the effect of substrate concentration on carbon isotope fractionation and the importance of monitoring isotope fractionation during the course of acetate consumption.     

Temperature Dependence of Carbon Isotope Fractionation in CAM Plants

The carbon isotope fractionation associated with nocturnal malic acid synthesis in Kalanchoë daigremontiana and Bryophyllum tubiflorum was calculated from the isotopic composition of carbon-4 of malic acid, after appropriate corrections. In the lowest temperature treatment (17°C nights, 23°C days), the isotope fractionation for both plants is −4‰ (that is, malate is enriched in ¹³C relative to the atmosphere). For K. daigremontiana, the isotope fractionation decreases with increasing temperature, becoming approximately 0‰ at 27°C/33°C. Detailed analysis of temperature effects on the isotope fractionation indicates that stomatal aperture decreases with increasing temperature and carboxylation capacity increases. For B. tubiflorum, the temperature dependence of the isotope fractionation is smaller and is principally attributed to the normal temperature dependences of the rates of diffusion and carboxylation steps. The small change in the isotopic composition of remaining malic acid in both species which is observed during deacidification indicates that malate release, rather than decarboxylation, is rate limiting in the deacidification process.     

Stable Hydrogen and Carbon Isotope Fractionation during Microbial Toluene Degradation: Mechanistic and Environmental Aspects

Primary features of hydrogen and carbon isotope fractionation during toluene degradation were studied to evaluate if analysis of isotope signatures can be used as a tool to monitor biodegradation in contaminated aquifers. D/H hydrogen isotope fractionation during microbial degradation of toluene was measured by gas chromatography. Per-deuterated toluene-d₈ and nonlabeled toluene were supplied in equal amounts as growth substrates, and kinetic isotope fractionation was calculated from the shift of the molar ratios of toluene-d₈ and nondeuterated toluene. The D/H isotope fractionation varied slightly for sulfate-reducing strain TRM1 (slope of curve [b] = −1.219), Desulfobacterium cetonicum (b = −1.196), Thauera aromatica (b = −0.816), and Geobacter metallireducens (b = −1.004) and was greater for the aerobic bacterium Pseudomonas putida mt-2 (b = −2.667). The D/H isotope fractionation was 3 orders of magnitude greater than the ¹³C/¹²C carbon isotope fractionation reported previously. Hydrogen isotope fractionation with nonlabeled toluene was 1.7 and 6 times less than isotope fractionation with per-deuterated toluene-d₈ and nonlabeled toluene for sulfate-reducing strain TRM1 (b = −0.728) and D. cetonicum (b = −0.198), respectively. Carbon and hydrogen isotope fractionation during toluene degradation by D. cetonicum remained constant over a growth temperature range of 15 to 37°C but varied slightly during degradation by P. putida mt-2, which showed maximum hydrogen isotope fractionation at 20°C (b = −4.086) and minimum fractionation at 35°C (b = −2.138). D/H isotope fractionation was observed only if the deuterium label was located at the methyl group of the toluene molecule which is the site of the initial enzymatic attack on the substrate by the bacterial strains investigated in this study. Use of ring-labeled toluene-d₅ in combination with nondeuterated toluene did not lead to significant D/H isotope fractionation. The activity of the first enzyme in the anaerobic toluene degradation pathway, benzylsuccinate synthase, was measured in cell extracts of D. cetonicum with an initial activity of 3.63 mU (mg of protein)⁻¹. The D/H isotope fractionation (b = −1.580) was 30% greater than that in growth experiments with D. cetonicum. Mass spectroscopic analysis of the product benzylsuccinate showed that H atoms abstracted from the toluene molecules by the enzyme were retained in the same molecules after the product was released. Our findings revealed that the use of deuterium-labeled toluene was appropriate for studying basic features of D/H isotope fractionation. Similar D/H fractionation factors for toluene degradation by anaerobic bacteria, the lack of significant temperature dependence, and the strong fractionation suggest that analysis of D/H fractionation can be used as a sensitive tool to assess degradation activities. Identification of the first enzyme reaction in the pathway as the major fractionating step provides a basis for linking observed isotope fractionation to biochemical reactions.     

Carbon and Hydrogen Isotopic Fractionation during Anaerobic Biodegradation of Benzene

Compound-specific isotope analysis has the potential to distinguish physical from biological attenuation processes in the subsurface. In this study, carbon and hydrogen isotopic fractionation effects during biodegradation of benzene under anaerobic conditions with different terminal-electron-accepting processes are reported for the first time. Different enrichment factors () for carbon (range of −1.9 to −3.6‰) and hydrogen (range of −29 to −79‰) fractionation were observed during biodegradation of benzene under nitrate-reducing, sulfate-reducing, and methanogenic conditions. These differences are not related to differences in initial biomass or in rates of biodegradation. Carbon isotopic enrichment factors for anaerobic benzene biodegradation in this study are comparable to those previously published for aerobic benzene biodegradation. In contrast, hydrogen enrichment factors determined for anaerobic benzene biodegradation are significantly larger than those previously published for benzene biodegradation under aerobic conditions. A fundamental difference in the previously proposed initial step of aerobic versus proposed anaerobic biodegradation pathways may account for these differences in hydrogen isotopic fractionation. Potentially, C-H bond breakage in the initial step of the anaerobic benzene biodegradation pathway may account for the large fractionation observed compared to that in aerobic benzene biodegradation. Despite some differences in reported enrichment factors between cultures with different terminal-electron-accepting processes, carbon and hydrogen isotope analysis has the potential to provide direct evidence of anaerobic biodegradation of benzene in the field.     

Mitochondrial Respiration Can Support NO(3) and NO(2) Reduction during Photosynthesis : Interactions between Photosynthesis, Respiration, and N Assimilation in the N-Limited Green Alga Selenastrum minutum

Mass spectrometric analysis shows that assimilation of inorganic nitrogen (NH₄⁺, NO₂⁻, NO₃⁻) by N-limited cells of Selenastrum minutum (Naeg.) Collins results in a stimulation of tricarboxylic acid cycle (TCA cycle) CO₂ release in both the light and dark. In a previous study we have shown that TCA cycle reductant generated during NH₄⁺ assimilation is oxidized via the cytochrome electron transport chain, resulting in an increase in respiratory O₂ consumption during photosynthesis (HG Weger, DG Birch, IR Elrifi, DH Turpin [1988] Plant Physiol 86: 688-692). NO₃⁻ and NO₂⁻ assimilation resulted in a larger stimulation of TCA cycle CO₂ release than did NH₄⁺, but a much smaller stimulation of mitochondrial O₂ consumption. NH₄⁺ assimilation was the same in the light and dark and insensitive to DCMU, but was 82% inhibited by anaerobiosis in both the light and dark. NO₃⁻ and NO₂⁻ assimilation rates were maximal in the light, but assimilation could proceed at substantial rates in the light in the presence of DCMU and in the dark. Unlike NH₄⁺, NO₃⁻ and NO₂⁻ assimilation were relatively insensitive to anaerobiosis. These results indicated that operation of the mitochondrial electron transport chain was not required to maintain TCA cycle activity during NO₃⁻ and NO₂⁻ assimilation, suggesting an alternative sink for TCA cycle generated reductant. Evaluation of changes in gross O₂ consumption during NO₃⁻ and NO₂⁻ assimilation suggest that TCA cycle reductant was exported to the chloroplast during photosynthesis and used to support NO₃⁻ and NO₂⁻ reduction.     

Carbon isotope ratios demonstrate carbon flux from c(4) host to c(3) parasite

Carbon isotope ratios of mature leaves from the C₃ angiosperm root hemiparasites Striga hermonthica (Del.) Benth (−26.7‰) and S. asiatica (L.) Kuntze (−25.6‰) were more negative than their C₄ host, sorghum (Sorghum bicolor [L.] Moench cv CSH1), (−13.5‰). However, in young photosynthetically incompetent plants of S. hermonthica this difference was reduced to less than 1‰. Differences between the carbon isotope ratios of two C₃-C₃ associations, S. gesnerioides (Willd.) Vatke—Vigna unguiculata (L.) Walp. and Oryza sativa L.—Rhamphicarpa fistulosa (Hochst.) Benth differed by less than 1‰. Theoretical carbon isotope ratios for mature leaves of S. hermonthica and S. asiatica, calculated from foliar gas exchange measurements, were −31.8 and −32.0‰, respectively. This difference between the measured and theoretical δ¹³C-values of 5 to 6‰ suggests that even in mature, photosynthetically active plants, there is substantial input of carbon from the C₄ host. We estimate this to be approximately 28% of the total carbon in S. hermonthica and 35% in S. asiatica. This level of carbon transfer contributes to the host's growth reductions observed in Striga-infected sorghum.     

Microbial Isotopic Fractionation of Perchlorate Chlorine

Perchlorate contamination can be microbially respired to innocuous chloride and thus can be treated effectively. However, monitoring a bioremediative strategy is often difficult due to the complexities of environmental samples. Here we demonstrate that microbial respiration of perchlorate results in a significant fractionation (~−15‰) of the chlorine stable isotope composition of perchlorate. This can be used to quantify the extent of biotic degradation and to separate biotic from abiotic attenuation of this contaminant.     

Variation in the carbon isotope composition of a plant with crassulacean Acid metabolism

The content of ¹³C varies in plants with Crassulacean acid metabolism. Differences up to 3.5‰ in the ¹³C/¹²C ratios were observed between leaves of different age in the same plant of Bryophyllum daigremontianum. Soluble and insoluble carbon in the same leaf differed up to 8‰, the largest difference occurring in the leaves with the highest Crassulacean acid metabolism activity. Models to account for the isotope discrimination by C₃, C₄, and Crassulacean acid metabolism plants are proposed.     

Photosynthetic carbon metabolism of a marine grass

The δ¹³C value of a tropical marine grass Thalassia testudinum is −9.04‰. This value is similar to the δ¹³C value of terrestrial tropical grasses. The δ¹³C values of the organic acid fraction, the amino acid fraction, the sugar fraction, malic acid, and glucose are: −11.2‰, −13.1‰, −10.1‰, −11.1‰, and −11.5‰, respectively. The δ¹³C values of malic acid and glucose of Thalassia are similar to the δ¹³C values of these intermediates in sorghum leaves and attest to the presence of the photosynthetic C₄-dicarboxylic acid pathway in this marine grass. The inorganic HCO₃⁻ for the growth of the grass fluctuates between −6.7 to −2.7‰ during the day. If CO₂ fixation in Thalassia is catalyzed by phosphoenolpyruvate carboxylase (which would result in a −3‰ fractionation between HCO₃⁻ and malic acid), the predicted δ¹³C value for Thalassia would be −9.7 to −5.7‰. This range is close to the observed range of −12.6 to −7.8‰ for Thalassia and agree with the operation of the C₄-dicarboxylic acid pathway in this plant. The early products of the fixation of HCO₃⁻ in the leaf sections are malic acid and aspartic acid which are similar to the early products of CO₂ fixation in C₄ terrestrial plants.     

Stable Carbon Isotope Fractionation by Sulfate-Reducing Bacteria

Biogeochemical transformations occurring in the anoxic zones of stratified sedimentary microbial communities can profoundly influence the isotopic and organic signatures preserved in the fossil record. Accordingly, we have determined carbon isotope discrimination that is associated with both heterotrophic and lithotrophic growth of pure cultures of sulfate-reducing bacteria (SRB). For heterotrophic-growth experiments, substrate consumption was monitored to completion. Sealed vessels containing SRB cultures were harvested at different time intervals, and δ¹³C values were determined for gaseous CO₂, organic substrates, and products such as biomass. For three of the four SRB, carbon isotope effects between the substrates, acetate or lactate and CO₂, and the cell biomass were small, ranging from 0 to 2‰. However, for Desulfotomaculum acetoxidans, the carbon incorporated into biomass was isotopically heavier than the available substrates by 8 to 9‰. SRB grown lithoautotrophically consumed less than 3% of the available CO₂ and exhibited substantial discrimination (calculated as isotope fractionation factors [α]), as follows: for Desulfobacterium autotrophicum, α values ranged from 1.0100 to 1.0123; for Desulfobacter hydrogenophilus, the α value was 0.0138, and for Desulfotomaculum acetoxidans, the α value was 1.0310. Mixotrophic growth of Desulfovibrio desulfuricans on acetate and CO₂ resulted in biomass with a δ¹³C composition intermediate to that of the substrates. The extent of fractionation depended on which enzymatic pathways were used, the direction in which the pathways operated, and the growth rate, but fractionation was not dependent on the growth phase. To the extent that environmental conditions affect the availability of organic substrates (e.g., acetate) and reducing power (e.g., H₂), ecological forces can also influence carbon isotope discrimination by SRB.     

Isotopic Incorporation Rates and Discrimination Factors in Mantis Shrimp Crustaceans

Stable isotope analysis has provided insights into the trophic ecology of a wide diversity of animals. Knowledge about isotopic incorporation rates and isotopic discrimination between the consumer and its diet for different tissue types is essential for interpreting stable isotope data, but these parameters remain understudied in many animal taxa and particularly in aquatic invertebrates. We performed a 292-day diet shift experiment on 92 individuals of the predatory mantis shrimp, Neogonodactylus bredini, to quantify carbon and nitrogen incorporation rates and isotope discrimination factors in muscle and hemolymph tissues. Average isotopic discrimination factors between mantis shrimp muscle and the new diet were 3.0 ± 0.6 ‰ and 0.9 ± 0.3 ‰ for carbon and nitrogen, respectively, which is contrary to what is seen in many other animals (e.g. C and N discrimination is generally 0–1 ‰ and 3–4 ‰, respectively). Surprisingly, the average residence time of nitrogen in hemolymph (28.9 ± 8.3 days) was over 8 times longer than that of carbon (3.4 ± 1.4 days). In muscle, the average residence times of carbon and nitrogen were of the same magnitude (89.3 ± 44.4 and 72.8 ± 18.8 days, respectively). We compared the mantis shrimps’ incorporation rates, along with rates from four other invertebrate taxa from the literature, to those predicted by an allometric equation relating carbon incorporation rate to body mass that was developed for teleost fishes and sharks. The rate of carbon incorporation into muscle was consistent with rates predicted by this equation. Our findings provide new insight into isotopic discrimination factors and incorporation rates in invertebrates with the former showing a different trend than what is commonly observed in other animals.     

Carbon Isotope Fractionation during Aerobic Biodegradation of Trichloroethene by Burkholderia cepacia G4: a Tool To Map Degradation Mechanisms

The strain Burkholderia cepacia G4 aerobically mineralized trichloroethene (TCE) to CO₂ over a time period of ~20 h. Three biodegradation experiments were conducted with different bacterial optical densities at 540 nm (OD₅₄₀s) in order to test whether isotope fractionation was consistent. The resulting TCE degradation was 93, 83.8, and 57.2% (i.e., 7.0, 16.2, and 42.8% TCE remaining) at OD₅₄₀s of 2.0, 1.1, and 0.6, respectively. ODs also correlated linearly with zero-order degradation rates (1.99, 1.11, and 0.64 μmol h⁻¹). While initial nonequilibrium mass losses of TCE produced only minor carbon isotope shifts (expressed in per mille δ¹³C_VPDB), they were 57.2, 39.6, and 17.0‰ between the initial and final TCE levels for the three experiments, in decreasing order of their OD₅₄₀s. Despite these strong isotope shifts, we found a largely uniform isotope fractionation. The latter is expressed with a Rayleigh enrichment factor, , and was −18.2 when all experiments were grouped to a common point of 42.8% TCE remaining. Although, decreases of to −20.7 were observed near complete degradation, our enrichment factors were significantly more negative than those reported for anaerobic dehalogenation of TCE. This indicates typical isotope fractionation for specific enzymatic mechanisms that can help to differentiate between degradation pathways.     

Enzymatic fractionation of carbon isotopes by phosphoenolpyruvate carboxylase from c(4) plants

The carbon atoms of glucose and malate in C₄ plants are 2 to 3‰ enriched in ¹²C with respect to atmospheric CO₂; whereas these intermediates in C₃ plants are 15 to 18‰ enriched with ¹²C with respect to atmospheric CO₂. The enzymatic synthesis of malate from phosphoenolpyruvate and bicarbonate in preparations of leaves of Sorghum bicolor, Haygrazer result in a carbon isotope fractionation of about 3‰. The enzymatic synthesis of phosphoglyceric acid from ribulose 1,5-diP and CO₂ in these preparations (contaminated with carbonic anhydrase) at 24 C and 37 C result in a carbon isotope fractionation of 33.7‰ and 18.3‰, respectively. These data are consistent with the conclusion that the small enrichment of ¹²C in the carbon atoms of malate and glucose (with respect to atmospheric CO₂) in leaves of Sorghum bicolor, Haygrazer occurs at the phosphoenolpyruvate carboxylase step.     

C/C ratio changes in crassulacean Acid metabolism plants

¹³C/¹²C ratios have been found in totally combusted leaves of Crassulacean acid metabolism plants to range from −14 to −33 δ ¹³C‰ compared with a limestone standard. Crassulacean acid metabolism plants apparently utilize both ribulose-1, 5-diphosphate carboxylase and phosphoenolpyruvate carboxylase to assimilate atmospheric CO₂ and, depending on environmental conditions, have ¹³C/¹²C ratios indicative of either carboxylase or to any intermediate value. The degree of discrimination against ¹³C and the resultant ¹³C/¹²C ratio from the photosynthetically fixed CO₂ is influenced by environmental conditions and is not a specific and fixed characteristic of a Crassulacean acid metabolism plant. Certain Crassulacean acid metabolism plants may shift their ratios as much as 17 δ ¹³C‰ in specific environments.     

Chaetognatha of the Namibian Upwelling Region: Taxonomy,Distribution and Trophic Position

In October 2010, the vertical distribution, biodiversity and maturity stages of Chaetognatha species were investigated at four stations located off Walvis Bay, Namibia. Seventeen species were detected and classified as pelagic, shallow-mesopelagic, deep-mesopelagic and bathypelagic species based upon the weighted mean depth derived from their average vertical distribution. High abundances of Chaetognatha were found in the upper 100 m at all stations of the Walvis Bay transect with a maximum value of 20837 ind. 1000 m⁻³ at the outer shelf station near the surface. The community was dominated by species of the Serratodentata group. Furthermore, the distribution of Chaetognatha did not seem to be influenced by low oxygen concentrations. Stable isotope ratios of carbon and nitrogen in Chaetognatha were determined for seven different areas located off northern Namibia. The values of δ¹⁵N ranged from 6.05 ‰ to 11.39 ‰, while the δ¹³C values varied between −23.89 ‰ and −17.03 ‰. The highest values for δ¹⁵N were observed at the Walvis Bay shelf break station. The lowest δ¹³C values were found at the Rocky Point offshore station, which was statistically different from all other areas. Stable isotopes of carbon and nitrogen were determined for four taxa (Sagitta minima, Planctonis group, Sagitta enflata, Sagitta decipiens). In this case, the δ¹⁵N values ranged from 6.17 ‰ to 10.38 ‰, whereas the δ¹³C values varied from −22.70 ‰ to −21.56 ‰. The lowest δ¹⁵N values were found for S. minima. The C- and N-content revealed maximum C-values for S. decipiens and maximum N-values for the Planctonis group. The C:N ratio of Chaetognatha ranged between 5.25 and 6.20. Overall, Chaetognatha are a diverse group in the pelagic food web of the Benguela Upwelling System and act as competitors of fish larvae and jelly fish by preying on copepods.