Automated system for simultaneous analysis of delta(13)C, delta(18)O and CO(2) concentrations in small air samples

In this paper we present an automated system for simultaneous measurement of CO(2) concentration, delta(13)C and delta(18)O from small (<1 mL) air samples in a short period of time (approximately 1 hour). This system combines continuous-flow isotope ratio mass spectrometry (CF-IRMS) and gas chromatography (GC) with an inlet system similar to conventional dual-inlet methods permitting several measurement cycles of standard and sample air. Analogous to the dual-inlet method, the precision of this system increases with the number of replicate cycles measured. The standard error of the mean for a measurement with this system is 0.7 ppm for the CO(2) concentration and 0.05 per thousand for the delta(13)C and delta(18)O with four replicate cycles and 0.4 ppm and 0.03 per thousand respectively with nine replicate cycles. The mean offset of our measurements from NOAA/CMDL analyzed air samples was 0.08 ppm for the CO(2) concentration, 0.01 per thousand for delta(13)C and 0.00 per thousand for delta(18)O. A specific list of the parts and operation of the system is detailed as well as some of the applications for micrometeorological and ecophysiological applications.     

Continuous flow stable isotope methods for study of delta(13)C fractionation during halomethane production and degradation

Gas chromatography/mass spectrometry/isotope ratio mass spectrometry (GC/MS/IRMS) methods for delta(13)C measurement of the halomethanes CH(3)Cl, CH(3)Br, CH(3)I and methanethiol (CH(3)SH) during studies of their biological production, biological degradation, and abiotic reactions are presented. Optimisation of gas chromatographic parameters allowed the identification and quantification of CO(2), O(2), CH(3)Cl, CH(3)Br, CH(3)I and CH(3)SH from a single sample, and also the concurrent measurement of delta(13)C for each of the halomethanes and methanethiol. Precision of delta(13)C measurements for halomethane standards decreased (+/-0.3, +/-0.5 and +/-1.3 per thousand) with increasing mass (CH(3)Cl, CH(3)Br, CH(3)I, respectively). Given that carbon isotope effects during biological production, biological degradation and some chemical (abiotic) reactions can be as much as 100 per thousand, stable isotope analysis offers a precise method to study the global sources and sinks of these halogenated compounds that are of considerable importance to our understanding of stratospheric ozone destruction.     

Multiple stable isotope (18O, 13C, 15N and 34S) analysis of human hair to identify the recent migrants in a rural community in SW England

Relationships between recent migration and hair delta(18)O values were examined for 40 people living in a rural community in SW England. The isotopic contents of 35 'local' hair samples were compared with those of 5 recently arrived individuals (from Australia, Canada, Chile, Germany and the USA). The hair delta(18)O values of these 'visitors' were +7.9 (Omaha, USA), +11.2 (Jena, Germany), +12.1 (Osorno, Chile), +12.6 (Montreal, Canada) and +14.3 per thousand (Adelaide, Australia). The hair value for the USA visitor (+7.9 per thousand) fell outside the range for the 33 local adult residents, +10.5 to +14.3 per thousand (+12.7 +/- 0.8 per thousand). Hair delta(18)O values did not identify the individuals from Adelaide, Montreal and Osorno as 'visitors', but hair delta(13)C or delta(34)S data did. Combining the hair delta(18)O, delta(13)C and delta(34)S values using principal components analysis (two components explained 89% of the overall variation among the 40 subjects) helped to more clearly distinguish European from non-European individuals, indicating the existence of global overall isotope (geo-origin) relationships.     

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.     

Validating methods for measuring delta18O and delta13C in otoliths from freshwater fish

The ability of the phosphoric acid digestion technique to extract carbon dioxide from biogenic carbonates and reliably reproduce delta(18)O and delta(13)C signatures from standard reference materials (NBS-18, NBS-19) was tested and shown to produce accurate, unbiased measurements of non-biologic materials. The effects of roasting preparation methods commonly reported when analyzing biogenic carbonates were also tested in a series of experiments using reference standards and otoliths obtained from aquacultured Arctic charr and rainbow trout. Roasting had no effect on the isotope measurement of reference standards. No significant differences between mean oxygen isotope signatures from paired experiments with roasted and non-roasted fish otoliths were found. However, otolith oxygen isotope measurements were significantly enriched in comparison to rearing water-based measurements for both species. Agreement between expected isotopic equilibrium and measured otolith delta(18)O values varied as a function of roasting temperature and between species. Criteria for the selection of appropriate roasting temperatures are suggested and favour 350 degrees C in freshwater fish where unbiased estimates of average rearing water temperatures and known differences in rearing temperatures were obtained. Carbon isotopic disequilibria were observed for both species. A mixing model analysis established differences in the percentage of metabolically derived carbon in studied otoliths, with Arctic charr deriving a greater proportion of otolith delta(13)C from metabolism as a result of higher metabolic rates.     

(18)O/(16)O isotope effect on miscibility of isobutyric acid with water

The miscibility of isobutyric acid with water and the influence of the isotope substitution of oxygen (16O/18O) in water over a broad concentration range are reported. The system exhibits a phase diagram with an upper critical solution temperature (UCST) and a visible isotope effect thereon. The oxygen isotope substitution decreases the UCST, leading to better miscibility of isobutyric acid with water. The isotope shift, DeltaTc = Tc(18O) - Tc(16O), extrapolated to the 100% enrichment in 18O, is equal to -1.15 K; hence it is much smaller and opposite to that found for deuterium substitution. The origin of the observed miscibility isotope effect has been qualitatively discussed in terms of the condensed-phase isotope effect theory.     

Grain-scale heterogeneities in the stable carbon and oxygen isotopic compositions of the international standard calcite materials (NBS 19, NBS 18, IAEA-CO-1, and IAEA-CO-8)

We determined grain-scale heterogeneities (from 6 to 88 microg) in the stable carbon and oxygen isotopic compositions (delta(13)C and delta(18)O) of the international standard calcite materials (NBS 19, NBS 18, IAEA-CO-1, and IAEA-CO-8) using a continuous-flow isotope ratio mass spectrometry (CF-IRMS) system that realizes a simultaneous determination of the delta(13)C and the delta(18)O values with standard deviations (S.D.) of less than 0.05 per thousand for CO(2) gas. Based on the S.D. of the delta(13)C and delta(18)O values determined for CO(2) gases evolved from the different grains of the same calcite material, we found that NBS 19, IAEA-CO-1, and IEAE-CO-8 were homogeneous for delta(13)C (less than 0.10 per thousand S.D.), and that only NBS 19 was homogeneous for delta(18)O (less than 0.14 per thousand S.D.). On the level of single grains, we found that both IAEA-CO-1 and IAEA-CO-8 were heterogeneous for delta(18)O (1.46 per thousand and 0.76 per thousand S.D., respectively), and that NBS 18 was heterogeneous for both delta(13)C and delta(18)O (0.34 per thousand and 0.54 per thousand S.D., respectively). Closer inspection of NBS 18 grains revealed that the highly deviated isotopic compositions were limited to the colored grains. By excluding such colored grains, we could also obtain the homogeneous delta(13)C and delta(18)O values (less than 0.18 per thousand and less than 0.16 per thousand S.D., respectively) for NBS 18. We conclude that NBS 19, IAEA-CO-1, or pure grains in NBS 18 are suitable to be used as the standard reference material for delta(13)C, and that either NBS 19 or pure grains in NBS 18 are suitable to be used as the reference material for delta(18)O during the grain-scale isotopic analyses of calcite.     

δ13C and δ18O trends across overstory environments in whole foliage and cellulose of three Pinus species

Stable isotope ratios of carbon (delta(13)C) and oxygen (delta(18)O) are increasingly used to investigate environmental influences on plant physiology. Cellulose is often isolated for isotopic studies, but some authors have questioned the value of this process. We studied trends in delta(13)C and delta(18)O of whole foliage and holocellulose from seedlings of three Pinus species across three overstory environments to evaluate the benefits of holocellulose extraction in the context of a traditional ecological experiment. Both tissue types showed increasing delta(13)C from closed-canopy controls to thinned plots to 0.3 ha canopy gaps, and no change in delta(18)O between overstory environments. delta(13)C of P. resinosa and P. strobus was greater than delta(13)C of P. banksiana in whole foliage and holocellulose samples, and there were no differences in delta(18)O associated with species in either tissue type. Our results suggest whole foliage and holocellulose provide similar information about isotopic trends across broad environmental gradients and between species, but holocellulose may be better suited for studying differences in stable isotope composition between multiple species across several treatments.     

Probing adsorption sites of silica-supported platinum with (13)C(16)O + (12)C(16)O and (13)C(18)O + (12)C(16)O mixtures: a comparative fourier transform infrared investigation

A comparative investigation of the adsorption of (13)C(18)O + (12)C(16)O and (13)C(16)O + (12)C(16)O mixtures on silica-supported Pt has been conducted. It is advantageous to use (13)C(18)O + (12)C(16)O mixtures rather than (13)C(16)O + (12)C(16)O to probe the adsorption sites and electronic state of supported Group VIII metals because the vibrational bands of the adsorbed (13)C(18)O and (12)C(16)O isotopic molecules do not overlap. In addition, while an intensity redistribution suppresses the lower-frequency band with adsorbed (13)C(16)O and (12)C(16)O with vibrational frequencies differing by 50 cm(-1), the intensity redistribution is less pronounced with the adsorbed (13)C(18)O and (12)C(16)O in which the frequency difference is 100 cm(-1). Moreover, the small intensity redistribution that does occur between the bands of adsorbed (13)C(18)O and (12)C(16)O still allows the detection of the vibrational band of adsorbed (13)C(18)O at (13)C(18)O gas-phase concentrations as low as 3%. At such low concentrations, the dipole-dipole interaction between adsorbed (13)C(18)O molecules is negligible, and, hence, both the singleton frequency and the dipole-dipole shift for adsorbed CO may be obtained in a single experiment. Two types of strongly bound and one type of weakly bound linear CO-Pt adsorption complexes have been identified and characterized by their singleton frequencies and dipole-dipole coupling shifts. The origin of these CO adsorption modes is discussed.     

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.     

(13)C natural abundance in the British diet: implications for (13)C breath tests

Surprisingly little information is available on the natural abundance of the minor isotope of carbon, (13)C, in common foodstuffs in the British diet. This study therefore aimed to examine the (13)C natural abundance of foodstuffs from a small cross-section of the British diet. The isotopic abundance, delta per mil, was calculated by measurement of the isotope ratio (13)C:(12)C by isotope ratio mass spectrometry. Results from this study were also compared with results from a North American study to highlight the difference in isotopic abundance between Northern European foodstuffs and North American foodstuffs. Such data should prove useful to those planning tracer studies using the stable isotope (13)C where enrichment is measured against a large and variable natural abundance in the body. Minimisation of this basal variation, for example in breath CO(2), can be achieved by controlling dietary intake of foods naturally abundant in (13)C.     

Determination of hydrogen, carbon and oxygen isotope ratios of ethanol in aqueous solution at millimole levels

Three stable isotope ratios, D/H, (13)C/(12)C and (18)O/(16)O, are measurable in ethanol, an important organic compound that is used as a material for food and beverages, fuel and chemical feedstock, and as a substance related to metabolism. We developed a simple and rapid method of measurement of three isotope ratios of ethanol in aqueous solution at millimole levels using gas chromatography-high-temperature conversion or combustion-isotope ratio mass spectrometry (GC-TC/C-IRMS) combined with solid-phase microextraction (SPME). Using this method, the delta value for ethanol was determined in 30 min for deltaD and delta(13)C, and in 75 min for delta(18)O with precisions of +/-9 per thousand, +/-0.3 per thousand and +/-0.7 per thousand, respectively, for deltaD, delta(13)C, and delta(18)O. An advantage of this process is that it requires no distillation for ethanol purification. The method is useful for small quantities of analyte with low ethanol concentrations, which is expected for environmental and metabolic studies.     

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.     

Origin assessment of green coffee (Coffea arabica) by multi-element stable isotope analysis of caffeine

The delta(13)C(VPDB), delta(2)H(VSMOW) and delta(18)O(VSMOW) values of caffeine isolated from Arabica green coffee beans of different geographical origin have been determined by isotope ratio mass spectrometry (IRMS) using elemental analysis (EA) in the "combustion" (C) and "pyrolysis" (P) modes (EA-C/P-IRMS). In total, 45 coffee samples (20 from Central and South America, 16 from Africa, six from Indonesia, and three from Jamaica and Hawaii) were analysed, as well as three reference samples of synthetic caffeine. Validation was performed by excluding isotope discrimination in the course of sample preparation and determining linear dynamic ranges for EA-P-IRMS measurements. The values for caffeine from green coffee ranged from -25.1 to - 29.9 per thousand, -109 to -198 per thousand, and +2.0 to -12.0 per thousand for delta(13)C(VPDB), delta(2)H(VSMOW), and delta(18)O(VSMOW), respectively. Data evaluation by linear discrimination analysis (LDA) and by classification and regression tree (CART) analysis revealed the delta(18)O(VSMOW) values to be highly significant. Use of LDA on the delta(2)H(VSMOW) and delta(18)O(VSMOW) data from coffee of African and Central/South American provenance led to error rates of 5.7% and 7.7% for adaption and cross validation, respectively.     

High-precision determination of 18O/16O ratios of silver phosphate by EA-pyrolysis-IRMS continuous flow technique

A high-precision, and rapid on-line method for oxygen isotope analysis of silver phosphate is presented. The technique uses high-temperature elemental analyzer (EA)-pyrolysis interfaced in continuous flow (CF) mode to an isotopic ratio mass spectrometer (IRMS). Calibration curves were generated by synthesizing silver phosphate with a 13 per thousand spread in delta(18)O values. Calibration materials were obtained by reacting dissolved potassium dihydrogen phosphate (KH(2)PO(4)) with water samples of various oxygen isotope compositions at 373 K. Validity of the method was tested by comparing the on-line results with those obtained by classical off-line sample preparation and dual inlet isotope measurement. In addition, silver phosphate precipitates were prepared from a collection of biogenic apatites with known delta(18)O values ranging from 12.8 to 29.9 per thousand (V-SMOW). Reproducibility of +/- 0.2 per thousand was obtained by the EA-Py-CF-IRMS method for sample sizes in the range 400-500 microg. Both natural and synthetic samples are remarkably well correlated with conventional (18)O/(16)O determinations. Silver phosphate is a very stable material and easy to degas and, thus, could be considered as a good candidate to become a reference material for the determination of (18)O/(16)O ratios of phosphate by high-temperature pyrolysis.     

(13)C/(12)C isotope labelling to study leaf carbon respiration and allocation in twigs of field-grown beech trees

In situ (13)C/(12)C isotopic labelling was conducted in field-grown beech (Fagus sylvatica) twigs to study carbon respiration and allocation. This was achieved with a portable gas-exchange open system coupled to an external chamber. This method allowed us to subject leafy twigs to CO(2) with a constant carbon isotope composition (delta(13)C of -51.2 per thousand) in an open system in the field. The labelling was done during the whole light period at two different dates (in June 2002 and October 2003). The delta(13)C values of respiratory metabolites and CO(2) that is subsequently respired during the night were measured. It was found that night-respired CO(2) is not completely labelled (only ca. 58% and 27% of new carbon is found in respired CO(2) immediately after the labelling in June 2002 and October 2003, respectively) and the labelling level progressively disappeared during the next day. It is concluded that the carbon respired by beech leaves after illumination was supplied by a mixture of carbon sources in which current carbohydrates were not the only contributors. In addition, as has been found in herbaceous plants, isotopic data before labelling showed that carbon isotope discrimination favoring the (13)C isotope occurred during the night respiration of beech leaves.     

Influence of dietary composition on the carbon, nitrogen, oxygen and hydrogen stable isotope ratios of milk

The stable isotope ratios ((13)C/(12)C, (15)N/(14)N, (18)O/(16)O, D/H) of animal feed and milk were investigated, considering cows stabled in two farms and fed with diets made up of different kinds of C(3) plants and different amounts of maize. Maize was characterised by delta(13)C, delta(18)O and deltaD values significantly higher than those of the C(3) plants, while, for the C(3) plants, Festuca arudinacea had significantly higher content of (13)C and (15)N. The delta(13)C and delta(18)O values of the overall diet and the delta(13)C of milk casein and lipids were shown to be significantly correlated with the percentage of maize in the animal diet. On the other hand, the delta(18)O values of milk water and the delta(18)O, deltaD and delta(15)N values of casein were shown to be only slightly influenced by the amount of maize in the feed, being probably more closely correlated with the geo-climatic and pedological characteristics of the area of origin and with the presence of fresh plant or silage in the ration. The delta(13)C value of casein was shown to be a suitable parameter for evaluating the amount of maize in the diet: each 10% increase in the maize content corresponded to a shift of 0.7 per thousand to 1.0 per thousand in the delta(13)C of casein. A threshold value of -23.5 per thousand for delta(13)C in milk casein, above which it is not possible to exclude the presence of maize in the diet, was suggested. The results obtained could be useful for determining mislabelling of dairy products declared to have been produced by pastured animals or of PDO cheeses with an established amount of maize in the diet and for verifying the unpermitted addition of exogenous components to milk.     

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.     

Determination of (13)C natural abundance of amino acid enantiomers in soil: methodological considerations and first results

The application of a combined gas chromatography-combustion/isotope ratio mass spectrometry (GC-C/IRMS) method for stable carbon isotope analysis of amino acid enantiomers in soil samples is presented. Triplicate delta(13)C analyses of pentafluoropropionyl (PFP) isopropyl ester derivatives of 27 amino acid enantiomers revealed that discrimination of (13)C during derivatization is different for different amino acid enantiomers and different amounts. Injection of increasing amounts of amino acid derivatives showed that the isotopic signal varied up to 10 per thousand for D-aspartic acid. Correction for the delta(13)C signal of underivatized amino acid enantiomers is possible for all investigated amino acid enantiomers using logarithmic functions. Operating the GC-C/IRMS system in the split-mode (split ratio 1:12) is possible but resulted in a higher isotopic discrimination. The detection limit approached 3 ng for some amino acid enantiomers in the splitless mode, while the lower limit of routine determination exceeded 10 ng injection amount. The upper limit at which accurate stable isotope values were obtained was 200 ng injection amount. Compound-specific delta(13)C analysis of alanine, valine, aspartic and glutamic acid showed that the D-forms were enriched in (13)C relative to the L-forms, suggesting that microbes significantly contributed to the formation of the D-enantiomers in soil.     

Stable carbon and oxygen isotopic determination of sub-microgram quantities of CaCO3 to analyze individual foraminiferal shells

We have developed an analytical system to determine stable isotopic compositions (delta13C and delta18O) of sub-microgram quantities of CaCO3 for the purpose of analyzing individual foraminiferal shells, using continuous-flow isotope ratio mass spectrometry (CF-IRMS). The system consists of a micro-volume CaCO3 decomposition tube, stainless steel CO2 purification vacuum line with a quantity-regulating unit, helium-purged CO2 purification line, gas chromatograph, and a CF-IRMS system. By using this system, we can determine stable carbon and oxygen isotopic compositions as low as 0.2 microg of CaCO3, with standard deviations of +/-0.10 per thousand for delta13C and +/-0.18 per thousand for delta18O within a 4-h reaction time and 30-min analysis period.