Sample vial influences on the accuracy and precision of carbon and oxygen isotope ratio analysis in continuous flow mass spectrometric applications

Due to the small amounts of sample gas involved in continuous flow mass spectrometric analysis, care should be taken to evaluate the influence of sample containers on the carbon and oxygen isotope ratios of samples. Data indicate that Na-glass and borosilicate glass vials, equipped with butyl rubber septa, can cause significant changes in the isotopic composition of CO(2) gas, even where sample gases are stored within the vial for less than one day. The magnitude of these changes varies from vial to vial. Given the leverage that contamination can potentially exert on small gas samples, each researcher should carefully evaluate the effect of sample vials in order to eliminate unknown and unwanted changes in the composition of samples. Copyright 2000 John Wiley & Sons, Ltd.     

Analysis of low‐concentration gas samples with continuous‐flow isotope ratio mass spectrometry: eliminating sources of contamination to achieve high precision

Developments in continuous‐flow isotope ratio mass spectrometry have made possible the rapid analysis of δ¹³C in CO₂ of small‐volume gas samples with precisions of ≤0.1‰. Prior research has validated the integrity of septum‐capped vials for collection and short‐term storage of gas samples. However, there has been little investigation into the sources of contamination during the preparation and analysis of low‐concentration gas samples. In this study we determined (1) sources of contamination on a Gasbench II, (2) developed an analytical procedure to reduce contamination, and (3) identified an efficient, precise method for introducing sample gas into vials. We investigated three vial‐filling procedures: (1) automated flush‐fill (AFF), (2) vacuum back‐fill (VBF), and (3) hand‐fill (HF). Treatments were evaluated based on the time required for preparation, observed contamination, and multi‐vial precision. The worst‐case observed contamination was 4.5% of sample volume. Our empirical estimate showed that this level of contamination results in an error of 1.7‰ for samples with near‐ambient CO₂ concentrations and isotopic values that followed a high‐concentration carbonate reference with an isotope ratio of ?47‰ (IAEA‐CO‐9). This carry‐over contamination on the Gasbench can be reduced by placing a helium‐filled vial between the standard and the succeeding sample or by ignoring the first two of five sample peaks generated by each analysis. High‐precision (SD ≤0.1‰) results with no detectable room‐air contamination were observed for AFF and VBF treatments. In contrast, the precision of HF treatments was lower (SD ≥0.2‰). VBF was optimal for the preparation of gas samples, as it yielded faster throughput at similar precision to AFF. Copyright © 2009 John Wiley & Sons, Ltd.     

Collection and storage of CO2 for 13C analysis: An application to separate soil CO2 efflux into root- and soil-derived components

Soil surface CO2 efflux is comprised of CO2 from (i) root respiration and rhizosphere microbes and (ii) heterotrophic respiration from the breakdown of soil organic matter (SOM). This efflux may be partitioned between these sources using delta13C measurements. To achieve this, continuous flow isotope ratio mass spectrometry can be used and, in conjunction with 10 mL septum-capped vials, large numbers of samples may be analysed using a Finnigan MAT Delta(plus)XP interfaced to a Gas Bench II. Here we describe a number of advances to facilitate such work, including: (i) a technique for monitoring mass spectrometer performance, (ii) improvements to sample storage, and (iii) a gas-handling system for incubating and sampling the CO2 derived from roots and soils. Mass spectrometer performance was monitored using an automated refillable vial. Compressed air analysed with this system had mean delta13C of -9.61 +/- 0.16 per thousand (+/- 1sigma, n = 28) collected over four runs. Heating the butyl rubber septa used to seal the vials at 105 degrees C for 12 h improved the sample storage. After air transportation over 12 days, the isotope composition of the CO2 at ambient concentrations was unchanged (before: -35.2 +/- 0.10 per thousand, n = 4; after: -35.3 +/- 0.10 per thousand, n = 15); without heat treatment of the septa the CO2 became slightly enriched (-35.0 +/- 0.14 per thousand, n = 15). The linearity of the Gas Bench II was found to decline above 8000 micromol CO2 mol(-1). To stay within a linear range and to allow the incubation of soil and root material we describe a gas-handling system based around a peristaltic pump. Finally, we demonstrate these methods by growing a C-4 grass (Guinea grass, Panicum maximum Jacq.) in a C-3 soil. Root respiration was found to contribute between 5 and 22% to the soil surface CO2 efflux. These methodologies will facilitate experiments aimed at measuring the isotopic composition of soil-derived CO2 across a range of ecological applications.     

Extraction of CO2 from air samples for isotopic analysis and limits to ultra high precision delta18O determination in CO2 gas.

The determination of delta18O values in CO2 at a precision level of +/-0.02 per thousand (delta-notation) has always been a challenging, if not impossible, analytical task. Here, we demonstrate that beyond the usually assumed major cause of uncertainty - water contamination - there are other, hitherto underestimated sources of contamination and processes which can alter the oxygen isotope composition of CO2. Active surfaces in the preparation line with which CO2 comes into contact, as well as traces of air in the sample, can alter the apparent delta18O value both temporarily and permanently. We investigated the effects of different surface materials including electropolished stainless steel, Duran glass, gold and quartz, the latter both untreated and silanized. CO2 frozen with liquid nitrogen showed a transient alteration of the 18O/16O ratio on all surfaces tested. The time to recover from the alteration as well as the size of the alteration varied with surface type. Quartz that had been ultrasonically cleaned for several hours with high purity water (0.05 microS) exhibited the smallest effect on the measured oxygen isotopic composition of CO2 before and after freezing. However, quartz proved to be mechanically unstable with time when subjected to repeated large temperature changes during operation. After several days of operation the gas released from the freezing step contained progressively increasing trace amounts of O2 probably originating from inclusions within the quartz, which precludes the use of quartz for cryogenically trapping CO2. Stainless steel or gold proved to be suitable materials after proper pre-treatment. To ensure a high trapping efficiency of CO2 from a flow of gas, a cold trap design was chosen comprising a thin wall 1/4" outer tube and a 1/8" inner tube, made respectively from electropolished stainless steel and gold. Due to a considerable 18O specific isotope effect during the release of CO2 from the cold surface, the thawing time had to be as long as 20 min for high precision delta18O measurements. The presence of traces of air in almost all CO2 gases that we analyzed was another major source of error. Nitrogen and oxygen in the ion source of our mass spectrometer (MAT 252, Finnigan MAT, Bremen, Germany) give rise to the production of NO2 at the hot tungsten filament. NO2+ is isobaric with C16O18O+ (m/z 46) and interferes with the delta18O measurement. Trace amounts of air are present in CO2 extracted cryogenically from air at -196 degrees C. This air, trapped at the cold surface, cannot be pumped away quantitatively. The amount of air present depends on the surface structure and, hence, the alteration of the measured delta18O value varies with the surface conditions. For automated high precision measurement of the isotopic composition of CO2 of air samples stored in glass flasks an extraction interface ('BGC-AirTrap') was developed which allows 18 analyses (including standards) per day to be made. For our reference CO2-in-air, stored in high pressure cylinders, the long term (>9 months) single sample precision was 0.012 per thousand for delta13C and 0.019 per thousand for delta18O.     

Carbon-isotopic composition of soil-respired carbon dioxide in static closed chambers at equilibrium

The carbon-isotopic composition (delta13C) of soil-respired CO2 has been employed to evaluate soil carbon-cycling processes and the contribution of soil CO2 emissions to canopy and tropospheric air. These evaluations can be successful only when accurate isotope values of soil-respired CO2 are available. Here, we tested the robustness of delta13C values of soil-respired CO2 obtained after long incubations in static closed chambers that were initially flushed with soil air. The rationale of this approach is that the equilibrium carbon-isotope values of chamber-headspace CO2 are theoretically equal to those of CO2 produced within the soil. Static closed chambers were installed in replicated grass monocultures, and measurements of headspace CO2 concentrations and delta13C values were performed at regular time intervals for 24 h in July 2005. The results revealed no significant effects of grass species on headspace CO2 concentrations or delta13C values (repeated measures analysis of variance (ANOVA), P>0.1). As predicted by theory, isotope values asymptotically approached equilibrium conditions, which in our experimental setting occurred after 10 h. This good match between model predictions and our results suggests that an accurate determination of delta13C values of CO2 produced within soils is obtained through the isotopic measurement of chamber-headspace CO2 once equilibrium conditions have been reached with the underlying soils. An additional advantage of this approach is that only one sample per chamber is required, which, combined with the low uncertainties of these measurements, facilitates the investigation of the spatial (landscape) variability of soil-respired CO2.     

Evaluating high time-resolved changes in carbon isotope ratio of respired CO2 by a rapid in-tube incubation technique

Recent insights into fractionation during dark respiration and rapid dynamics in isotope signatures of leaf- and ecosystem-respired CO(2) indicate the need for new methods for high time-resolved measurements of the isotopic signature of respired CO(2) (delta(13)C(res)). We present a rapid and simple method to analyse delta(13)C(res) using an in-tube incubation technique and an autosampler for small septum-capped vials. The effect of storage on the delta(18)O and delta(13)C ratios of ambient CO(2) concentrations was tested with different humidity and temperatures. delta(13)C ratios remained stable over 72 h, whereas delta(18)O ratios decreased after 24 h. Storage at 4 degrees C improved the storage time for delta(18)O. Leaves or leaf discs were incubated in the vials, flushed with CO(2)-free air and respired CO(2) was automatically sampled within 5 min on a microGas autosampler interfaced to a GV-Isoprime isotope ratio mass spectrometer. Results were validated by simultaneous on-line gas-exchange measurements of delta(13)C(res) of attached leaves. This method was used to evaluate the short-term (5-60 min) and diurnal dynamics of delta(13)C(res) in an evergreen oak (Quercus ilex) and a herb (Tolpis barbata). An immediate depletion of 2-4 per thousand from the initial delta(13)C(res) value occurred during the first 30 min of darkening. Q. ilex exhibited further a substantial diurnal enrichment in delta(13)C(res) of 8 per thousand, followed by a progressive depletion during the night. In contrast, T. barbata did not exhibit a distinct diurnal pattern. This is in accordance with recent theory on fractionation in metabolic pathways and may be related to the different utilisation of the respiratory substrate in the fast-growing herb and the evergreen oak. These data indicate substantial and rapid dynamics (within minutes to hours) in delta(13)C(res), which differed between species and probably the growth status of the plant. The in-tube incubation method enables both high time-resolved analysis and extensive sampling across different organs, species and functional types.     

Stable carbon and oxygen isotopic analysis of carbon monoxide in natural waters

Techniques have been developed to allow on-line simultaneous analysis of concentration and stable isotopic compositions ((13)C and (18)O) of dissolved carbon monoxide (CO) in natural water, using continuous-flow isotope ratio mass spectrometry (CF-IRMS). The analytical system consisted sequentially of a He-sparging bottle of water, a gas dryer, CO(2)-trapping stage using both Ascarite trap and silica-gel packed gas chromatography (GC), on-line oxidation to CO(2) using the Schütze reagent, cryofocusing, GC purification using a capillary column and measurement by CF-IRMS. Each sample analysis takes about 40 minutes. The detection limit with delta(13)C standard deviation of 0.5 per thousand is 300 pmol and that with delta(18)O deviation of 1.0 per thousand is 750 pmol. Analytical blanks associated with these methods are 21+/-9 pmol. The procedures are evaluated through analyses of temporally varying concentration and isotopic compositions of CO in an artificial lake on the university campus. The delta(13)C and delta(18)O values of CO showed wide variation in accordance with diurnal variation of CO concentration, probably due to significant isotopic effects during photochemical production and microbial oxidation of CO in the aquatic environment. The delta(13)C and delta(18)O values of CO should be a useful tool in studies of the mechanism and pathways of CO production and consumption in natural waters.     

A rapid and precise technique for measuring delta(13)C-CO(2) and delta(18)O-CO(2) ratios at ambient CO(2) concentrations for biological applications and the influence of container type and storage time on the sample isotope ratios

A simple modification to a commercially available gas chromatograph isotope ratio mass spectrometer (GC/IRMS) allows rapid and precise determination of the stable isotopes ((13)C and (18)O) of CO(2) at ambient CO(2) concentrations. A sample loop was inserted downstream of the GC injection port and used to introduce small volumes of air samples into the GC/IRMS. This procedure does not require a cryofocusing step and significantly reduces the analysis time. The precisions for delta(13)C and delta(18)O of CO(2) at ambient concentration were +/-0.164 and +/-0.247 per thousand, respectively. This modified GC/IRMS was used to test the effects of storage on the (18)O and (13)C isotopic ratios of CO(2) at ambient concentrations in four container types. On average, the change in the (13)C-CO(2) and (18)O-CO(2) ratios of samples after one week of storage in glass vials equipped with butyl rubber stoppers (Bellco Glass Inc.) were depleted by 0.12 and by 0.20 per thousand, respectively. The (13)C ratios in aluminum canisters (Scotty II and IV, Scott Specialty Gasses) after one month of storage were depleted, on average, by 0.73 and 2.04 per thousand, respectively, while the (18)O ratios were depleted by 0.38 and 1.20 per thousand for the Scotty II and IV, respectively. After a month of storage in electropolished containers (Summa canisters, Biospheric Research Corporation), the (13)C-CO(2) and (18)O-CO(2) ratios were depleted, on average, by 0.26 and enriched by 0.30 per thousand, respectively, close to the precision of measurements. Samples were collected at a mature hardwood forest for CO(2) concentration determination and isotopic analysis. A comparison of CO(2) concentrations determined with an infrared gas analyzer and from sample voltages, determined on the GC/IRMS concurrent with the isotopic analysis, indicated that CO(2) concentrations can be determined reliably with the GC/IRMS technique. The (13)C and (18)O ratios of nighttime ecosystem-respired CO(2), determined from the intercept of Keeling plots, were -26.11 per thousand (V-PDB) and -8.81 per thousand (V-PDB-CO(2)), respectively.     

Optimized sample preparation for isotopic analyses of CO2 in air: systematic study of precision and accuracy dependence on driving variables during CO2 purification process

A systematic analysis of efficiency, reproducibility and accuracy of cryogenic purification of CO(2) from air samples for isotopic analyses is presented. The technical characteristics of the cryogenic line are given in detail. To study the cryogenic process, three different operating parameters are considered: flow rate of the gas entering the line, pressure of the gas in the line, and CO(2)-trap shape. Experimental results demonstrate that efficiency, reproducibility and accuracy strongly depend on the CO(2)trap shape. Moreover, a dependence of reproducibility and accuracy on the flow rate of the gas is found, but not on its pressure. High precision (< or =0.02 per thousand for delta(13)C and < or =0.05 per thousand for delta(18)O) and good accuracy (<0.09 per thousand for delta(13)C and <0.14 per thousand for delta(18)O) is achieved after applying the N(2)O correction.     

A field and laboratory method for monitoring the concentration and isotopic composition of soil CO2

The stable isotope composition of nmol size gas samples can be determined accurately and precisely using continuous flow isotope ratio mass spectrometry (IRMS). We have developed a technique that exploits this capability in order to measure delta13C and delta18O values and, simultaneously, the concentration of CO2 in sub-mL volume soil air samples. A sampling strategy designed for monitoring CO2 profiles at particular locations of interest is also described. This combined field and laboratory technique provides several advantages over those previously reported: (1) the small sample size required allows soil air to be sampled at a high spatial resolution, (2) the field setup minimizes sampling times and does not require powered equipment, (3) the analytical method avoids the introduction of air (including O2) into the mass spectrometer thereby extending filament life, and (4) pCO2, delta13C and delta18O are determined simultaneously. The reproducibility of measurements of CO2 in synthetic tank air using this technique is: +/-0.08 per thousand (delta13C), +/-0.10 per thousand (delta18O), and +/-0.7% (pCO2) at 5550 ppm. The reproducibility for CO2 in soil air is estimated as: +/-0.06 per thousand (delta13C), +/-0.06 per thousand (delta18O), and +/-1.6% (pCO2). Monitoring soil CO2 using this technique is applicable to studies concerning soil respiration and ecosystem gas exchange, the effect of elevated atmospheric CO2 (e.g. free air carbon dioxide enrichment) on soil processes, soil water budgets including partitioning evaporation from transpiration, pedogenesis and weathering, diffuse solid-earth degassing, and the calibration of speleothem and pedogenic carbonate delta13C values as paleoenvironmental proxies.     

A gas chromatography/combustion/isotope ratio mass spectrometry system for high-precision delta13C measurements of atmospheric methane extracted from ice core samples

Past atmospheric composition can be reconstructed by the analysis of air enclosures in polar ice cores which archive ancient air in decadal to centennial resolution. Due to the different carbon isotopic signatures of different methane sources high-precision measurements of delta13CH4 in ice cores provide clues about the global methane cycle in the past. We developed a highly automated (continuous-flow) gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) technique for ice core samples of approximately 200 g. The methane is melt-extracted using a purge-and-trap method, then separated from the main air constituents, combusted and measured as CO2 by a conventional isotope ratio mass spectrometer. One CO2 working standard, one CH4 and two air reference gases are used to identify potential sources of isotope fractionation within the entire sample preparation process and to enhance the stability, reproducibility and accuracy of the measurement. After correction for gravitational fractionation, pre-industrial air samples from Greenland ice (1831 +/- 40 years) show a delta13C(VPDB) of -49.54 +/- 0.13 per thousand and Antarctic samples (1530 +/- 25 years) show a delta13C(VPDB) of -48.00 +/- 0.12 per thousand in good agreement with published data.     

A simple method for oxygen-18 determination of milligram quantities of water using NaHCO3 reagent

This paper presents a modified H(2)O-CO(2) equilibration method for stable oxygen isotopic composition (delta(18)O) analysis of water. This method enables rapid and simple delta(18)O analysis of milligram quantities of water, by employing solid reagent NaHCO(3) as the CO(2) source, a small (0.6 mL) glass vial for the equilibration chamber, and an isotope-monitoring gas chromatography/mass spectrometry (irm-GC/MS) system for delta 18O(CO2) analysis. This method has several advantages, including simple handling for the H(2)O-CO(2) equilibration (without purging and/or evacuation treatments), rapid and easy delta(18)O analysis of equilibrated CO(2), and highly sensitive and highly precise delta(18)O analysis of H(2)O, using samples as small as 10 mg and with a precision of less than +/-0.12 per thousand. The time needed to attain oxygen isotopic equilibration between CO(2) and water is also comparable (17 h for 10 mg H(2)O and 10 h for 100 mg H(2)O) to other previous methods using CO(2) gas for the CO(2) source. The extent of delta(18)O variation of sample water from its initial delta(18)O value due to isotope exchange with added NaHCO(3) is also discussed. It is concluded that the correction needed is negligible (less than 0.1 per thousand ) as long as the oxygen atom ratio (O(NaHCO3)/O(H2O)) is less than 3.3 +/- 10(-3) and provided the delta18O(H2O) determination is made by comparing delta(18)O of CO(2) equilibrated with sample water and that equilibrated with standard water of a moderately close delta(18)O value, less than 30 per thousand difference.     

Assessing the stable carbon isotopic composition of intercellular CO2 in a CAM plant using gas chromatography-combustion-isotope ratio mass spectrometry

Most of the literature focused on internal CO(2) (Ci) determinations in plants has used indirect methods based on gas-exchange estimations. We have developed a new method based on the capture of internal air gas samples and their analysis by gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). This method provided a direct measure of intercellular CO(2) concentrations combined with stable carbon isotopic composition in O. ficus-indica plants. Plants were grown at both ambient and elevated CO(2) concentration. During the day period, when the stomata are closed, the Ci was high and was very (13)C-enriched in both ambient and elevated CO(2)-grown plants, reflecting Rubisco's fractionation (this plant enzyme has been shown to discriminate by 29 per thousand, in vitro, against (13)CO(2)). Other enzyme fractionations involved in C metabolism in plants, such as carbonic anhydrase, could also be playing an important role in the diurnal delta(13)C enrichment of the Ci. During the night, when stomata are open, Ci concentrations were higher in elevated (and the corresponding delta(13)C values were more (13)C-depleted) than in ambient CO(2)-grown plants.     

A portable automated system for trace gas sampling in the field and stable isotope analysis in the laboratory

A computer-controllable mobile system is presented which enables the automatic collection of 33 air samples in the field and the subsequent analysis for delta13C and delta18O stable isotope ratios of a carbon-containing trace gas in the laboratory, e.g. CO2, CO or CH4. The system includes a manifold gas source input for profile sampling and an infrared gas analyzer for in situ CO2 concentration measurements. Measurements of delta13C and delta18O of all 33 samples can run unattended and take less than six hours for CO2. Laboratory tests with three gases (compressed air with different pCO2 and stable isotope compositions) showed a measurement precision of 0.03 per thousand for delta13C and 0.02 per thousand for delta18O of CO2 (standard error (SE), n = 11). A field test of our system, in which 66 air samples were collected within a 24-hour period above grassland, showed a correlation of 0.99 (r2) between the inverse of pCO2 and delta13C of CO2. Storage of samples until analysis is possible for about 1 week; this can be an important factor for sampling in remote areas. A wider range of applications in the field is open with our system, since sampling and analysis of CO and CH4 for stable isotope composition is also possible. Samples of compressed air had a measurement precision (SE, n = 33) of 0.03 per thousand for delta13C and of 0.04 per thousand for delta18O on CO and of 0.07 per thousand for delta13C on CH4. Our system should therefore further facilitate research of trace gases in the context of the carbon cycle in the field, and opens many other possible applications with carbon- and possibly non-carbon-containing trace gases.     

Site-specific carbon isotope analysis of aromatic carboxylic acids by elemental analysis/pyrolysis/isotope ratio mass spectrometry

Site-specific carbon isotope composition of organic compounds can provide useful information on their origin and history in natural environments. Site-specific isotope analyses of small amounts of organic compounds (sub-nanomolar level), such as short-chain carboxylic acids and amino acid analogues, have been performed using gas chromatography/pyrolysis/isotope ratio mass spectrometry (GC/pyrolysis/IRMS). These analyses were previously limited to volatile compounds. In this study, site-specific carbon isotope analysis has been developed for non-volatile aromatic carboxylic acids at sub-micromolar level by decarboxylation using a continuous flow elemental analysis (EA)/pyrolysis/IRMS technique. Benzoic acid, 2-naphthylacetic acid and 1-pyrenecarboxylic acid were pyrolyzed at 500-1000 degrees C by EA/pyrolysis/IRMS to produce CO2 for delta13C measurement of the carboxyl group. These three aromatic acids were most efficiently pyrolyzed at 750 degrees C. Conventional sealed-tube pyrolysis was also conducted for comparison. The delta13C values of CO2 generated by the continuous flow technique were within 1.0 per thousand of those performed by the conventional technique, indicating that the new continuous flow technique can accurately analyze the carbon isotopic composition of the carboxyl group in aromatic carboxylic acids. The new continuous flow technique is simple, rapid and uses small sample sizes, so this technique will be useful for characterizing the isotopic signature of carboxyl groups in organic compounds.     

Assessing the use of delta(13)C natural abundance in separation of root and microbial respiration in a Danish beech (Fagus sylvatica L.) forest

Our understanding of forest biosphere-atmosphere interactions is fundamental for predicting forest ecosystem responses to climatic changes. Currently, however, our knowledge is incomplete partly due to inability to separate the major components of soil CO(2) effluxes, viz. root respiration, microbial decomposition of soil organic matter and microbial decomposition of litter material. In this study we examined whether the delta(13)C characteristics of solid organic matter and respired CO(2) from different soil-C components and root respiration in a Danish beech forest were useful to provide information on the root respiration contribution to total CO(2) effluxes. The delta(13)C isotopic analyses of CO(2) were performed using a FinniganMAT Delta(PLUS) isotope-ratio mass spectrometer coupled in continuous flow mode to a trace gas preparation-concentration unit (PreCon). Gas samples in 2-mL crimp seal vials were analysed in a fully automatic mode with an experimental standard error +/-0.11 per thousand. We observed that the CO(2) derived from root-free mineral soil horizons (A, B(W)) was more enriched in (13)C (delta(13)C range -21.6 to -21.2 per thousand ) compared with CO(2) derived from root-free humus layers (delta(13)C range -23.6 to -23.4 per thousand ). The CO(2) evolved from root respiration in isolated young beech plants revealed a value intermediate between those for the soil humus and mineral horizons, delta(13)C(root) = -22.2 per thousand, but was associated with great variability (SE +/- 1.0 per thousand ) due to plant-specific differences. delta(13)C of CO(2) from in situ below-ground respiration averaged -22.8 per thousand, intermediate between the values for the humus layer and root respiration, but variability was great (SE +/- 0.4 per thousand ) due to pronounced spatial patterns. Overall, we were unable to statistically separate the CO(2) of root respiration vs. soil organic matter decomposition based solely on delta(13)C signatures, yet the trend in the data suggests that root respiration contributed approximately 43% to total respiration. The vertical gradient in delta(13)C, however, might be a useful tool in partitioning respiration in different soil layers. The experiment also showed an unexpected (13)C-enrichment of CO(2) (>3.5 per thousand ) compared with the total-C signatures in the individual soil-C components. This may suggest that analyses of bulk samples are not representative for the C-pools actively undergoing decomposition.     

Mobile, outdoor continuous-flow isotope-ratio mass spectrometer system for automated high-frequency 13C- and 18O-CO2 analysis for Keeling plot applications

A continuous-flow isotope-ratio mass spectrometer (CF-IRMS, custom-made GasBenchII and Delta(plus)Advantage, ThermoFinnigan) was installed on a grassland site and interfaced with a closed-path infrared gas analyser (IRGA). The CF-IRMS and IRGA were housed in an air-conditioned travel van. Air was sampled at 1.5 m above the 0.07-m tall grassland canopy, drawn through a 17-m long PTFE tube at a rate of 0.25 L s(-1), and fed to the IRGA and CF-IRMS in series. The IRMS was interfaced with the IRGA via a stainless steel capillary inserted 0.5 m into the sample air outlet tube of the IRGA (forming an open split), a gas-tight pump, and a sample loop attached to the eight-port Valco valve of the continuous-flow interface. Air was pumped through the 0.25-mL sample loop at 10 mL s(-1) (a flushing frequency of 40 Hz). Air samples were analysed at intervals of approx. 2.8 min. Whole system precision was tested in the field using air mixed from pure CO2 and CO2-free air by means of mass flow controllers. The standard deviation of repeated single measurements was 0.21-0.07 per thousand for delta13C and 0.34-0.14 per thousand for delta18O of CO2 in air with mixing ratios ranging between 200-800 micromol mol(-1). The CO2 peak area measured by the IRMS was proportional to the CO2 mixing ratio (r2 = 1.00), allowing estimation of sample air CO2 mixing ratio from IRMS data. A 1-day long measurement cycle of CO2, delta13C and delta18O of air sampled above the grassland canopy was used to test the system for Keeling plot applications. Delta18O exhibited a clear diurnal cycle (4 per thousand range), but short-term (1-h interval) variability was small (average SD 0.38 per thousand). Yet, the correlation between delta18O and CO2 mixing ratio was relatively weak, and this was true for both the whole data set and 1-h subsets. Conversely, the delta13C of all 541 samples measured during the 25.2-h interval fitted well the Keeling regression (r2 = 0.99), yielding an intercept of -27.40 per thousand (+/-0.07 per thousand SE). Useful Keeling regressions (r2 > 0.9, average r2 = 0.96) also resulted from data collected over 1-h intervals of the 12-h long twilight and dark period. These indicated that 13C content of ecosystem respiration was approx. constant near -27.6 per thousand. The precision of the present system is similar to that of current techniques used in ecosystem studies which employ flask sampling and a laboratory-based CF-IRMS. Sampling (and measurement) frequency is greatly increased relative to systems based on flask sampling, and sampling time (0.025 s per sample) is decreased. These features increase the probability for sampling the entire CO2 range which occurs in a given time window. The system obviates sample storage problems, greatly minimises handling needs, and allows extended campaigns of high frequency sampling and analysis with minimal attendance.     

Methods to adjust for the interference of N2O on delta13C and delta18O measurements of CO2 from soil mineralization

In this paper we present an overview of the present knowledge relating to methods that avoid interference of N2O on delta13C and delta18O measurements of CO2. The main focus of research to date has been on atmospheric samples. However, N2O is predominantly generated by soil processes. Isotope analyses related to soil trace gas emissions are often performed with continuous flow isotope ratio mass spectrometers, which do not necessarily have the high precision needed for atmospheric research. However, it was shown by using laboratory and field samples that a correction to obtain reliable delta13C and delta18O values is also required for a commercial continuous flow isotope ratio mass spectrometer. The capillary gas chromatography column of the original equipment was changed to a packed Porapak Q column. This adaptation resulted in an improved accuracy and precision of delta13C (standard deviation(Ghent): from 0.2 to 0.08 per thousand; standard deviation(Lincoln): from 0.2 to 0.13 per thousand) of CO2 for N2O/CO2 ratios up to 0.1. For delta18O there was an improvement for the standard deviation measured at Ghent University (0.13 to 0.08 per thousand) but not for the measurements at Lincoln University (0.08 to 0.23 per thousand). The benefits of using the packed Porapak Q column compared with the theoretical correction method meant that samples were not limited to small N(2)O concentrations, they did not require an extra N2O concentration measurement, and measurements were independent of the variable isotopic composition of N2O from soil.     

A new method to determine the 17O isotopic abundance in CO2 using oxygen isotope exchange with a solid oxide.

This paper discusses a simple method to determine 17O isotope excess or deficiency ('mass-independent isotopic composition') in CO2 gas. When applying conventional mass spectrometry of CO2 (m/z 44, 45 and 46) to determine the 17O/16O ratio, the 13C/12C ratio has to be established separately. This can be achieved by analysing an aliquot of sample CO2 before and after subjecting it to oxygen isotope exchange with a pool of oxygen with 'normal' 17O/16O ratio, i.e. with Delta17O approximately equal to delta17O-0.516 x delta18O = 0. Cerium oxide has been shown to be practically well suited for the exchange of CO2 oxygen; the reagent is safe and does not produce any contamination. The CO2-CeO2 exchange reaction has 99.8 +/- 0.7% recovery yield. At 650 degrees C this reaction reaches equilibrium in 30 min and, as tested, results in complete oxygen replacement. Delta17O determinations depend on accuracy of CO2 delta measurements: the repeatability of +/-0.015 per thousand (1sigma) in delta(45)R and delta(46)R determination relative to the working reference results in an error of Delta17O as small as +/-0.33 per thousand. Such a precision is sufficient for Delta17O determination in stratospheric CO2. The calculated Delta17O value systematically depends on absolute 17R and 13R ratios in isotopic reference materials, which are presently not yet known with certainty (the 17R value is most important), and may be inadequate for 17O-correction with a = 0.516. Within the present uncertainty, Delta17O determined in 17O-enriched CO2 agrees with the value directly measured in the enriched O2 from which this CO2 was produced. Besides Delta17O determination, investigated CO2-CeO2 equilibration may have several other implications. Fast, complete isotopic exchange of CO2 by reaction with CeO2 may also be employed to get reproducible 17O-correction and, hence, to better monitor small delta13C shifts and to isotopically equilibrate mixtures of CO2 gases.     

Automated analysis of 13C/12C ratios in CO2 and dissolved inorganic carbon for ecological and environmental applications

Stable carbon isotope ratios (13C/12C) are a valuable tool for studying a wide range of environmental processes, including carbon cycling and subsurface microbial activity. Recent advances in automated analysis provide the opportunity to increase greatly the ease and consistency of isotopic analysis. This study evaluated an automated headspace sampler linked to a commercially available CO2 preconcentration system and continuous flow isotope ratio mass spectrometer. Field sampling and analysis methods are illustrated for delta13C of soil respired CO2, from both tracer and natural abundance experiments, and dissolved inorganic carbon from contaminated groundwater. The automated system demonstrated accuracy, precision, and linearity, with standard errors below 0.1 per thousand for replicate gas standards run at concentrations varying five-fold. It measured 40 samples per 10-hour run, with concentrations ranging from ppb to percentage levels. In the field, gas samples were injected into nitrogen-filled autosampler vials, thereby allowing use of small sample volumes, control of analyte concentration, and direct analysis by the automated system with no further preparation. A significant linear relationship between standard concentrations and peak area allows for accurate estimates of sample CO2 concentration from the mass spectrometric data. The ability to analyze multiple small-volume samples with minimal off-line preparation should enhance the application of isotopes to well-replicated field experiments for process-level studies and spatial and temporal scaling.