Determination Of Carbon Isotope Ratios Research Articles

Improving the Determination of Carbon Isotope Ratios of Endogenous Steroids Found in Human Serum.

The determination of serum concentrations of testosterone (T) and 4-androstenedione (A4) was implemented into the steroidal module of the Athlete Biological Passport in 2023. Monitoring T, A4, and the ratio of T/A4 in a longitudinal manner enables the detection of the misuse of low-dose T administrations especially in female athletes, whereas urinary markers of the steroid profile may not be influenced significantly. In contrast to the urinary steroid profile, knowledge on confounding factors regarding serum concentrations of T and A4 is yet comparably scarce, and corroborating exogenous sources of the target analytes by isotope ratio mass spectrometry (IRMS) is desirable. In a recent study, it was demonstrated that carbon isotope ratios (CIRs) of serum steroids can be determined if analyte concentrations permit. The therein-employed method utilized 2D-GC/IRMS, and only a limited number of potential endogenous reference compounds were available. The here-presented approach uses complementary analyte purification strategies, addressing previous limitations. A high-performance liquid chromatography cleanup was developed and fully validated for serum steroids in order to enable all doping control laboratories to potentially implement this method alongside existing protocols for urinary steroids. Besides the already-investigated endogenous steroids cholesterol, dehydroepiandrosterone sulfate, androsterone sulfate, and epiandrosterone sulfate, two additional steroids were included in the test menu, namely, pregnenolone sulfate and 5-androstene-3β,17β-diol sulfate. Serum steroid concentrations down to 25 ng/mL were found to allow robust CIR determinations, and a reference population encompassing 124 male and female athlete samples was investigated to enable the calculation of population-based thresholds for all relevant steroid combinations.

High-temperature liquid chromatography-isotope ratio mass spectrometry methodology for carbon isotope ratio determination of anabolic steroids in urine

BackgroundGas Chromatography Isotope Ratio Mass Spectrometry (GC-C-IRMS) has long been used in routine laboratories to determine the δ13C values of anabolic steroids in urine, differentiating between, e.g., endogenous and synthetic testosterone (T) in sports doping control. Until now, liquid chromatography (LC-IRMS) has not been used. The LC-IRMS setup doesn't allow organic solvents or modifiers in the mobile phase for δ13C determinations. Mid-to non-polar analytes such as steroids can be analysed in water heated to High Temperatures (HT, up to 200 °C) because at 200 °C has a similar polarity as 80/20 methanol/water at ambient temperature. In this work, we developed a method for steroids in urine, extending the application of the LC-IRMS to non-polar analytes in complex matrices. ResultAn HT-LC-IRMS method capable of determining the δ13C values of four steroids (i.e., testosterone (T), 5α-androstane-3α,17β-diol (ααβ), 5β-androstane-3α,17β-diol (βαβ) and pregnanetriol (PT)) in urine was developed and validated. Accuracy ranged from 0.23 ‰ (ααβ and βαβ) to 0.49 ‰ (T), and the detection limit was set at 10 ng mL−1 (T, ααβ+βαβ). The validation data and a comparison of authentic urine samples analysed with HT-LC-IRMS and GC-C-IRMS indicated a comparable performance between HT-LC-IRMS and GC-C-IRMS. SignificanceHT-LC-IRMS can be used to determine δ13C values of anabolic steroids, extending the applicability of both HT-LC and LC-IRMS to non-polar substances determined in a complex matrix in routine laboratory practice.

Employing 11-Ketotestosterone as a Target Analyte for Adrenosterone (11OXO) Administration in Doping Controls.

Adrenosterone (Androst-4-ene-3,11,17-trione, 11OXO) is forbidden in sports according to the Prohibited List of the World Anti-Doping Agency. The administration of 11OXO may be detected by monitoring the urinary concentrations of its main human metabolites 11β-hydroxy-androsterone and 11β-hydroxy-etiocholanolone. Preliminary urinary concentration and concentration ratio thresholds have been established for sports drug testing purposes, but adaptations are desirable as the suggested limits would result in numerous suspicious findings due to naturally elevated concentrations and ratios. Recently, the metabolism of 11-oxo-testosterone (KT) was investigated in the context of anti-doping research, resulting in a preliminary urinary concentration threshold and a confirmation procedure based on the determination of carbon isotope ratios (CIRs). Gas chromatography coupled to isotope ratio mass spectrometry was employed to investigate the CIRs of selected steroids. As KT is also a metabolite of 11OXO, the developed protocols for KT have been tested to elucidate their potential to detect the administration of 11OXO after a single oral dose of 100 mg. In order to further improve the analytical approach, the threshold for urinary concentrations of KT was re-investigated by employing a reference population of n = 5232 routine doping control samples. Quantification of urinary steroids was conducted by employing gas chromatography coupled to triple quadrupole mass spectrometry. Derived from these, a subset of n = 106 samples showing elevated concentrations of KT was investigated regarding their CIRs. By means of this, potentially positive samples due to the illicit administration of 11OXO or KT could be excluded, and the calculation of reference population-derived thresholds for the concentrations and CIR of KT was possible. Based on the results, the urinary concentration threshold for KT is suggested to be established at 130 ng/mL.

Investigations in carbon isotope ratios of seized testosterone and boldenone preparations.

In order to detect the misuse of testosterone (T) or boldenone (Bo) in doping control analysis, the confirmation of atypical findings employing the determination of carbon isotope ratios (CIR) is mandatory for issuing adverse analytical findings. Elevated concentrations of T (or elevated T/epitestosterone ratios) may result from confounding factors such as ethanol intake, and the presence of low urinary concentrations of Bo can originate from endogenous or urinary in situ production of small amounts of the steroid. As pharmaceutical preparations of Bo and T are generally depleted in 13 C, their CIR differ significantly from the 13 C-enriched endogenous steroids. Some rare cases have been reported on pharmaceutical preparations showing 13 C-enriched isotope ratios that complicate the current application of CIR in sports drug testing. Therefore, the CIR of a subset of n = 157 T preparations and n = 39 Bo preparations seized in Switzerland and Germany between 2013 and 2018 was analyzed in order to estimate the possible impact of steroid preparations showing 13 C-enriched isotope ratios on the current approach to detect their misuse. All investigated Bo preparations showed CIR in the expected range between - 26.7 and -30.3‰. Within the T samples, 95% showed the expected values below -26‰ while six samples fall between -25 and -26‰ and one sample was indistinguishable from endogenously produced T with a CIR of -23.3‰.

Analysis of endogenous steroids in urine by means of multi-immunoaffinity chromatography and isotope ratio mass spectrometry for sports drug testing.

Detecting the administration of naturally occurring but synthetically derived steroids (e.g., testosterone) in routine doping controls is particularly laborious and time-consuming. Carbon isotope signatures determined by isotope ratio mass spectrometry (IRMS) have been established as the method of choice to generate confirmatory evidence in case of suspicious or atypical findings in steroid profile analyses; however, IRMS measurements require sophisticated sample preparation methods employing up to two high-performance liquid chromatography (HPLC) purification steps. Here, an alternative sample preparation approach is presented. Immunoaffinity chromatography (IAC) was employed to reduce the batch analysis time by omitting the time-consuming HPLC purification steps, while pre- and post-IAC sample handling followed published protocols. IAC exploits specific antibody-immunogen interactions, and the option of combining three immunoaffinity gels containing specific antibodies for testosterone, pregnanediol, and 11-ketoetiocholanolone into a multi-immunoaffinity sample preparation approach was assessed. Due to cross reactivities, also etiocholanolone, androsterone, 5β-androstanediol, and 5α-androstanediol were co-extracted and included in the testing protocol. The method was validated by determining precision, recovery, and carry over, and performing linear mixing models. IAC was found to be applicable to the determination of carbon isotope ratios in doping controls and the approach allowed for an accelerated sample preparation. Graphical abstract.

Lessons learned from inter-laboratory studies of carbon isotope analysis of honey

Forensic application of carbon isotope ratio measurements of honey and honey protein to investigate the degree of adulteration with high fructose corn syrup or other C4 plant sugars is well established. These measurements must use methods that exhibit suitable performance criteria, particularly with regard to measurement uncertainty and traceability – low levels of adulteration can only be detected by methods that result in suitably small measurement uncertainties such that differences of 1‰ or less can be reliably detected. Inter-laboratory exercises are invaluable to assess the state-of-the art of measurement capabilities of laboratories necessary to achieve such performance criteria. National and designated metrology institutes from a number of countries recently participated in an inter-laboratory assessment (CCQM-K140) of stable carbon isotope ratio determination of bulk honey. The same sample material was distributed to a number of forensic isotope analysis laboratories that could not participate directly in the metrological comparison. The results from these studies have demonstrated that the majority of participants provided isotope delta values with acceptable performance metrics; that all participants ensured traceability of their results; and that where measurement uncertainties were reported; these were fit-for-purpose. A number of the forensic laboratories only reported precision rather than full estimates of measurement uncertainty and this was the major cause of the few instances of questionable performance metrics. Reporting of standard deviations in place of measurement uncertainties is common practice outside metrology institutes and the implications for interpretations of small differences in isotopic compositions are discussed. The results have also highlighted a number of considerations that are useful for organisers of similar inter-laboratory studies in the future.

Determination of carbon isotope ratios for honey samples by means of a liquid chromatography/isotope ratio mass spectrometry system coupled with a post-column pump.

Liquid chromatography/isotope ratio mass spectrometry (LC/IRMS) has been used to authenticate and trace products such as honey, wine, and lemon juice, and compounds such as caffeine and pesticides. However, LC/IRMS has several disadvantages, including the high cost of the CO2 membrane and blocking by solidified sodium persulfate. Here, we developed an improved system for determining carbon isotope ratios using LC/IRMS. The main improvement was the use of a post-column pump. Using the improved system, we determined δ13 C values for glucose with high accuracy and precision (0.1‰ and 0.1‰, respectively; n=3). The glucose, fructose, disaccharide, trisaccharide, and organic acid constituents of honey samples were analyzed using LC/IRMS. The δ13 C values for glucose, fructose, disaccharides, trisaccharides, and organic acids ranged from -27.0 to -24.2‰, -26.8 to -24.0‰, -28.8 to -24.0‰, -27.8 to -22.8‰, and-30.6 to -27.4‰, respectively. The analysis time was a third to a half of that required for analysis by previously reported methods. The column flow rate could be arbitrarily adjusted with the post-column pump. We applied the improved method to 26 commercial honey samples. Our results can be expected to be useful for other researchers who use LC/IRMS.

Authentication of citrus volatiles based on carbon isotope ratios

The theme of adulteration of natural flavours is briefly discussed along with the most common analytical tools used to unveil possible frauds of Citrus essential oils. Therefore, the article focuses on the main analytical approaches used for carbon isotope ratio determination of citrus secondary metabolites. Different applications of carbon isotope ratios mass spectrometry on the major Citrus products are revised to highlight the potentialities of this tool for authenticity assessment of Citrus essential oils and flavours. A deeper discussion is given on the most recent literature produced by the authors with a further evaluation of the complex picture emerged from their results. The data are overall considered to evaluate possible correlations among the carbon isotope ratios of different Citrus secondary metabolites and to prove whether univocal assessment of the genuineness of Citrus essential oils can be performed based on this analytical approach.

Development of methodology for the determination of carbon isotope ratios using gas chromatography/combustion/isotope ratio mass spectrometry and applications in the biodegradation of phenolic brominated flame retardants and their degradation products.

Compound-specific stable carbon isotope analysis of phenolic brominated flame retardants (BFRs) and bisphenol A (BPA) has proven informative to discriminate their source and fate apportionment in the environment. However, because these compounds contain highly polar functional groups and exist as complex mixtures in environment matrices, derivatization is a necessary step, which adds additional non-analyte carbon atoms for analyses and may alter the original stable carbon isotope ratio. It is, therefore, imperative to gain an insight into the relationship between the δ(13) C values of the BFRs and BPA derivatives and those of underivatized BFRs and BPA. The δ(13) C values of BFRs and BPA N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) derivatives were measured by gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS). The δ(13) C values of the BSTFA reagent and the standard underivatized BFRs and BPA were determined using elemental analyzer/isotope ratio mass spectrometry (EA/IRMS). The experimentally obtained δ(13) C values for BFRs and BPA derivatives were then compared with the theoretically calculated values. The derivatization process introduces no isotopic fractionation for BFRs and BPA (the average difference between the theoretically calculated and experimentally obtained δ(13) C values was 0.06 ± 0.15‰, within the precision limits of the GC/C/IRMS measurements). Therefore, the δ(13) C values for the original underivatized BFRs and BPA were computed through a mass balance equation. This work offers a novel tool to research the biotic or abiotic transformation processes of BFRs and BPA in the environment and will offer a perspective for the identification of the environmental source and fate of these organic compounds.

Validation of user- and environmentally friendly extraction and clean-up methods for compound-specific stable carbon isotope analysis of organochlorine pesticides and their metabolites in soils

In order to evaluate the potential of compound-specific stable carbon isotope analysis (CSIA) for tracking organochlorine pesticides in soil systems, sample pre-treatment methods have to be developed, which can provide recoveries sufficient for low detection limits without altering the isotope ratio of the target compounds. In this study we tested the compatibility of CSIA with user- and environmentally friendly extraction methods, including the Quick, Easy, Cheap, Effective, Rugged and Safe procedure (QuEChERS), Ultrasonic Assisted Extraction (USE) and Focused Ultrasonic Extraction (FUSE), as well as clean-up methods, including sulfuric acid clean-up and Florisil® column chromatography for hexachlorocyclohexanes (HCHs), p,pʹ-dichlorodiphenyltrichloroethane (DDT) and their environmental metabolites (chlorinated benzenes, dichlorodiphenyldichloroethylene – DDE and dichlorodiphenyldichloroethane – DDD) in soils. We optimized the extraction methods for maximum recovery and pre-concentration. At optimal conditions, all extraction methods and clean-up procedures, as well as the pre-concentration of the extract by solvent evaporation, led to insignificant changes in carbon isotope ratios of the target compounds. We modified the USE procedure to increase the volume of withdrawn organic extract, resulting in a higher pre-concentration of the target compounds by the subsequent solvent evaporation step. This Modified Ultrasonic Assisted Extraction (MUSE) was the most suitable procedure, and it was validated for the determination of carbon isotope ratios of the target compounds using two different types of soil matrices. The method could be applied to analyze carbon isotope ratios of HCHs, DDT, and their chlorinated metabolites in soil samples with concentrations ranging from 0.3 to 0.8mg/kg. The analytical uncertainty of MUSE, incorporating both accuracy and precision, was ≤0.4‰.

The emerging role of carbon isotope ratio determination in health research and medical diagnostics

Variations in the isotopic signature of carbon in biological samples (e.g. breath, blood and tissues) can be used to monitor shifts in whole body metabolism. As a conservative recorder of our diet, changes in the isotopic signature of carbon in biological samples provide an objective means to distinguish dietary patterns and the relationship with diseases. In addition, metabolic discrimination of carbon within the body can be informative regarding changes in the body's metabolic fuel usage during situations where shifts in the macronutrient oxidation ratio are expected. Therefore, changes in the isotopic signature over time have proven to be a tremendously powerful and sensitive means of detecting and measuring changes in steady-state systems. As such, this review focuses on how a naturally occurring ratio of stable isotopes of carbon (13C/12C) can be used as a biomarker for nutritional and metabolic status, altered macronutrient metabolism, and health and disease.

Position-specific isotope analysis of the methyl group carbon in methylcobalamin for the investigation of biomethylation processes

In the environment, the methylation of metal(loid)s is a widespread phenomenon, which enhances both biomobility as well as mostly the toxicity of the precursory metal(loid)s. Different reaction mechanisms have been proposed for arsenic, but not really proven yet. Here, carbon isotope analysis can foster our understanding of these processes, as the extent of the isotopic fractionation allows to differentiate between different types of reaction, such as concerted (SN2) or stepwise nucleophilic substitution (SN1) as well as to determine the origin of the methyl group. However, for the determination of the kinetic isotope effect the initial isotopic value of the transferred methyl group has to be determined. To that end, we used hydroiodic acid for abstraction of the methyl group from methylcobalamin (CH3Cob) or S-adenosyl methionine (SAM) and subsequent analysis of the formed methyl iodide by gas chromatography (GC) isotope ratio mass spectrometry (IRMS). In addition, three further independent methods have been investigated to determine the position-specific δ (13)C value of CH3Cob involving photolytic cleavage with different additives or thermolytic cleavage of the methyl-cobalt bonding and subsequent measurement of the formed methane by GC-IRMS. The thermolytic cleavage gave comparable results as the abstraction using HI. In contrast, photolysis led to an isotopic fractionation of about 7 to 9‰. Furthermore, we extended a recently developed method for the determination of carbon isotope ratios of organometal(loid)s in complex matrices using hydride generation for volatilization and matrix separation before heart-cut GC and IRMS to the analysis of the low boiling partly methylated arsenicals, which are formed in the course of arsenic methylation. Finally, we demonstrated the applicability of this methodology by investigation of carbon fractionation due to the methyl transfer from CH3Cob to arsenic induced by glutathione.

Carbon isotope ratio determination and investigation of seized testosterone preparations

In the present study, the content of a number of black market testosterone products collected in Austria has been analyzed. Additionally, (13) C/(12) C ratios were measured for testosterone in the products after cleavage of the testosterone ester. The aim was to determine whether some of these products had similar (13) C/(12) C ratios to those normally found for endogenous testosterone, which could prevent a positive isotopic ratio mass spectrometric (IRMS) finding in doping control. Moreover, it was investigated to what extent the preparations contained the masking agent epitestosterone, in order to lower the testosterone/epitestosterone (T/E) ratio in urinary steroid profiles. Out of 30 analyzed products, the declared ingredients differed from the actual content in 10 cases. Epitestosterone, however, could not be found in any of the products. The products displayed δ(13)C(VPDB) values between -23.6 and -29.4‰. For more than half of these products, the values were within a range reported for endogenous urinary steroids.

Determination of carbon isotope ratios of methanol and acetaldehyde in air samples by gas chromatography-isotope ratio mass spectrometry combined with headspace solid-phase microextraction

Isotopic signatures of atmospheric methanol and acetaldehyde have the potential to improve our ability to quantitatively assess their importance in atmospheric chemistry. However, isotopic measurements of atmospheric methanol and acetaldehyde and their individual source and sink processes have been limited. In this study, we examined gas chromatography-isotope ratio mass spectrometry combined with headspace solid-phase microextraction to measure the carbon isotope ratios of methanol and acetaldehyde in air samples. The method enabled us to determine carbon isotope ratios with a precision (1 standard deviation) of±0.6 ‰ for 20 ml of air sample containing more than 3 ppm of methanol and±0.7 ‰ for 20 ml of air sample containing more than 2 ppm of acetaldehyde. Moreover, the applicability of this method to determine isotope ratios of methanol and acetaldehyde emitted from detached plant leaves was demonstrated.

Surface Gas Geochemistry above the Natural CO2Reservoir of Montmiral (Drôme, France), Source Tracking and Gas Exchange between the Soil, Biosphere and Atmosphere

One of the options considered to mitigate greenhouse gas concentrations in the atmosphere is underground storage of CO<sub>2<sub/>. There is a strong need for enhancing and developing methods that would help throughout the duration life of such underground storage, to ensure the safety and able to monitor the evolution of the injected CO<sub>2<sub/> plume. Among these, geochemical methods can play an important role. Here, we describe results acquired under the research programme “Géocarbone-Monitoring”, partially funded by the French National Research Agency, on the Montmiral natural analogue in South-Eastern France. Other results obtained under the same research programme in the French Massif Central are reported elsewhere in this volume.Spot sampling methods allowing a great geographical coverage and continuous measurements on selected points were undertaken in 2006 and 2007, in order to determine soil gas concentrations and fluxes as well as carbon isotope ratio determinations. One important result is that without any evidence of deep CO<sub>2<sub/> leakage, both CO<sub>2<sub/> concentrations and fluxes appear to be higher than can be explained only by biological activities. Further investigations are thus needed to understand the gas evolution better throughout the year.

Precise and traceable carbon isotope ratio measurements by multicollector ICP-MS: what next?

This article reviews recent developments in the use of multicollector inductively coupled plasma mass spectrometry (MC-ICP-MS) to provide high-precision carbon isotope ratio measurements. MC-ICP-MS could become an alternative method to isotope ratio mass spectrometry (IRMS) for rapid carbon isotope ratio determinations in organic compounds and characterisation and certification of isotopic reference materials. In this overview, the advantages, drawbacks and potential of the method for future applications are critically discussed. Furthermore, suggestions for future improvements in terms of precision and sensitivity are made. No doubt, this is an exciting analytical challenge and, as such, hurdles will need to be cleared.

Flow Injection Analysis−Isotope Ratio Mass Spectrometry for Bulk Carbon Stable Isotope Analysis of Alcoholic Beverages

A new method for bulk carbon isotope ratio determination of water-soluble samples is presented that is based on flow injection analysis-isotope ratio mass spectrometry (FIA-IRMS) using an LC IsoLink interface. Advantages of the method are that (i) only very small amounts of sample are required (2-5 microL of the sample for up to 200 possible injections), (ii) it avoids complex sample preparation procedures such as needed for EA-IRMS analysis (only sample dilution and injection,) and (iii) high throughput due to short analysis times is possible (approximately 15 min for five replicates). The method was first tested and evaluated as a fast screening method with industrially produced ethanol samples, and additionally the applicability was tested by the measurement of 81 alcoholic beverages, for example, whiskey, brandy, vodka, tequila, and others. The minimal sample concentration required for precise and reproducible measurements was around 50 microL L(-1) ethanol/water (1.71 mM carbon). The limit of repeatability was determined to be r=0.49%. FIA-IRMS represents a fast screening method for beverage authenticity control. Due to this, samples can be prescreened as a decisive criterion for more detailed investigations by HPLC-IRMS or multielement GC-IRMS measurements for a verification of adulteration.

Determination of 13C/12C ratios of urinary epitestosterone and its main metabolites 5α‐ and 5β‐androstane‐3α, 17α‐diol

AbstractEpitestosterone (17α‐hydroxy‐androst‐4‐en‐3‐one, EpiT) belongs to the list of prohibited substances of the World Anti‐Doping Agency (WADA). Although it possesses no anabolic effect, it is presumed to be misused by athletes in order to mask administration of testosterone (T) by lowering the urinary T/EpiT ratio.To improve detection, an excretion study with 40 mg of orally administered EpiT was conducted focusing on the metabolites of EpiT: 5α‐ and 5β‐androstane‐3α,17 α‐diol (5aEpiD and 5bEpiD). A reference population of n = 74 volunteers was investigated to elucidate the urinary concentrations of these steroids.In order to prove whether an unusual finding in urinary concentrations or ratios is due to an illicit intake of steroids or due to physiological elevation, determination of carbon isotope ratios is advisable. A method for isotope ratio determination was developed to enable 13C/12C ratios of EpiT, 5bEpiD, 5aEpiD, pregnanediol and androsterone and etiocholanolone to be measured from a single urine specimen. The method's validity was tested by applying linear mixing models and specificity was ensured by means of gas chromatography/mass spectrometry analysis. δ13C values at natural levels were determined with a reference population and both Δ values and corresponding reference limits were calculated.Considering the implemented EpiT‐metabolites, a more than twofold extension of the detection time of EpiT administration was achieved with both the urinary concentration thresholds and the 13C/12C ratios. Copyright © 2009 John Wiley &amp; Sons, Ltd.

Membrane permeation continuous‐flow isotope ratio mass spectrometry for on‐line carbon isotope ratio determination

Gaseous membrane permeation (MP) technologies have been combined with continuous-flow isotope ratio mass spectrometry for on-line delta13C measurements. The experimental setup of membrane permeation-gas chromatography/combustion/isotope ratio mass spectrometry (MP-GC/C/IRMS) quantitatively traps gas streams in membrane permeation experiments under steady-state conditions and performs on-line gas transfer into a GC/C/IRMS system. A commercial polydimethylsiloxane (PDMS) membrane sheet was used for the experiments. Laboratory tests using CO2 demonstrate that the whole process does not fractionate the C isotopes of CO2. Moreover, the delta13C values of CO2 permeated on-line give the same isotopic results as off-line static dual-inlet IRMS delta13C measurements. Formaldehyde generated from aqueous formaldehyde solutions has also been used as the feed gas for permeation experiments and on-line delta13C determination. The feed-formaldehyde delta13C value was pre-determined by sampling the headspace of the thermostated aqueous formaldehyde solution. Comparison of the results obtained by headspace with those from direct aqueous formaldehyde injection confirms that the headspace sampling does not generate isotopic fractionation, but the permeated formaldehyde analyzed on-line yields a 13C enrichment relative to the feed delta13C value, the isotopic fractionation being 1.0026 +/- 0.0003. The delta13C values have been normalized using an adapted two-point isotopic calibration for delta13C values ranging from -42 to -10 per thousand. The MP-GC/C/IRMS system allows the delta13C determination of formaldehyde without chemical derivatization or additional analytical imprecision.

Chemical and isotopic analysis of hydrocarbon gas at trace levels: Methodology and results

Isotopic mass spectrometry coupled online with gas chromatography (GC-C-IRMS) permits measurement of relative proportions of gaseous hydrocarbon (CH 4 to C 4H 10) and CO 2, and determination of carbon isotope ratio of hydrocarbon molecules. Access to these parameters provides valuable information about the source and the genesis of naturally-occurring gas, as well as on post-formation physico-chemical processes which might have taken place in the geological environment. In particular, it is possible to distinguish hydrocarbon gas of bacterial origin from that of thermogenic origin based on proportion and carbon isotope ratio of methane as measured by GC-C-IRMS. However, in samples containing very low amounts of hydrocarbons (from 1 ppm to 1000 ppm), accurate measurement of isotope ratios is often impossible due to the limitations of conventional GC-C-IRMS techniques using direct sample introduction. A technique was developed to overcome this limitation. It is based on a novel approach allowing pre-concentration of hydrocarbons prior to GC-C-IRMS analysis. The pre-concentration step consists in selective trapping of hydrocarbon molecules on a cold adsorbent phase, and removal of non-adsorbed gases (N 2, O 2, Ar,…). In a second step, pre-concentrated alkanes are desorbed, and released in an inert carrier gas, focused through a capillary and introduced into the GC-C-IRMS for chromatographic separation and measurement of concentration and carbon isotope composition of each individual carbon molecule. In order to achieve sufficient accuracy, several operating conditions are of prime importance, including sufficient signal intensity, well defined peak shape and low signal/noise ratio. Accurate measurements can be performed on samples as small as 10 cm 3 of bulk gas in standard conditions, with concentrations as low as 1 ppm of methane, 0.5 ppm of ethane and 0.3 ppm of propane and butane. Total analytical uncertainty on δ 13C measurements ranges from ± 0.2‰ to ± 1.5‰, depending on the hydrocarbon molecule.