Relative Isotopic Abundances Research Articles

Nuclear hyperfine level pattern and hyperfine specific heat of frustrated Gd-pyrochlores Gd2 M 2O7 (M = Ti, Sn, Hf, Zr)

The nuclear contribution to the specific heat originates from the combination of a nuclear electric quadrupole interaction and a nuclear magnetic hyperfine interaction of the magnetic atoms embedded in the crystalline lattice structure. We have estimated the nuclear hyperfine level patterns of the nuclear ground state \(I_{g} = 3/2^{-}\) and the first excited state \(I_{ex} = 5/2^{+}\) at 86.5 keV of Mossbauer-active Gd155 nucleus of Gd-based pyrochlores Gd2M2O7 (M = Ti, Sn, Hf, Zr). The nuclear quadrupole and hyperfine interactions are scaled in accordance with the ratios of quadrupole and magnetic moments of \(I_{g} = 3/2^{-}\) state of Gd155 and Gd157 nuclei to get the contribution from Gd157 nucleus of Gd2M2O7 to the hyperfine specific heat Chf appropriate to its relative isotopic abundance. It is found that Chf(T) shows a Schottky hump around 6–8 mK for these pyrochlores. At temperatures much higher than the hyperfine level splitting, Chf behaves as AT − 2 with A ∼ 1–2 × 10 − 4 JK/mole-Gd which is consistent with experimental results.

Protein turnover: Measurement of proteome dynamics by whole animal metabolic labelling with stable isotope labelled amino acids

The measurement of protein turnover in tissues of intact animals is obtained by whole animal dynamic labelling studies, requiring dietary administration of precursor label. It is difficult to obtain full labelling of precursor amino acids in the diet and if partial labelling is used, calculation of the rate of turnover of each protein requires knowledge of the precursor relative isotope abundance (RIA). We describe an approach to dynamic labelling of proteins in the mouse with a commercial diet supplemented with a pure, deuterated essential amino acid. The pattern of isotopomer labelling can be used to recover the precursor RIA, and sampling of urinary secreted proteins can monitor the development of liver precursor RIA non-invasively. Time-series analysis of the labelling trajectories for individual proteins allows accurate determination of the first order rate constant for degradation. The acquisition of this parameter over multiple proteins permits turnover profiling of cellular proteins and comparisons of different tissues. The median rate of degradation of muscle protein is considerably lower than liver or kidney, with heart occupying an intermediate position.

A proteomics strategy for determining the synthesis and degradation rates of individual proteins in fish

In order to study the protein dynamics in the tissues of fish we have developed a proteomics-based strategy to determine the rates of synthesis and degradation of individual proteins. We have demonstrated the feasibility of this approach by measuring the turnover of multiple isoforms of parvalbumin (β1-7) in the skeletal muscle of common carp (Cyprinus carpio). A stable isotope-labelled amino acid ([(2)H(7)] l-leucine) was administered to the carp via the diet and its incorporation into the isoforms of parvalbumin in muscle over time was monitored by LC-MS analysis of signature peptides. The relative isotope abundance was calculated and used to deconvolute the data. The β7 parvalbumin isoform had a rate of synthesis that was greater than the rate of degradation. In contrast the rate of degradation of the β5 isoform exceeded its rate of synthesis, whilst the analysis revealed that the other parvalbumin β-isoforms (β1, β2, β3, β4 and β6) had a rate of synthesis that was equal to the rate of degradation. This work has addressed a number of technical challenges and represents the first study to use proteomic approaches to measure the turnover of individual proteins in fish.

Spectral accuracy of a new hybrid quadrupole time‐of‐flight mass spectrometer: application to ranking small molecule elemental compositions

Determining the elemental compositions of unknown molecules is an important goal of analytical chemistry. The isotope pattern revealed by a mass spectrometer provides valuable information regarding the elemental composition of a molecule. In order to employ spectral accuracy considerations for elemental composition determination, it is important to know how faithfully a mass spectrometer can record the isotope pattern and to understand the magnitude of the errors of the relative isotopic abundances. Twenty-four small molecule drugs and two natural products representing a diverse range of elemental compositions and ranging in molecular weight from 236 to 1663 Da were measured on a new hybrid orthogonal acceleration quadrupole time-of-flight (Q-TOF) mass spectrometer by flow infusion analysis. The similarity between the observed profile isotope pattern and the theoretical isotope pattern, denoted spectral accuracy, was calculated using a computational algorithm in the program MassWorks. The spectral accuracy for all compounds averaged better than 98%. When using spectral accuracy to rank elemental compositions with the elemental constraints (C(1-100)H(0-200)N(0-50)O(0-50)F(0-5)S(0-5)Cl(0-5)Br(0-5)) further restricted by empirical rules and a mass tolerance ≤5 parts-per-million, the correct formula was ranked first over 80% of the time. In contrast, when using mass accuracy for ranking, only two compounds (8%) were ranked first. For quinidine and troglitazone, the initial spectral accuracy measurements were lower than expected and further analysis indicated that minor, structurally related components were present. Our work has determined the magnitude of spectral accuracy that can be expected on a new Q-TOF mass spectrometer. In addition, we demonstrate the utility of spectral accuracy measurements both for ranking elemental compositions and also for obtaining insight into the chemical nature of the analyte that might otherwise be overlooked.

Accuracy of relative isotopic abundance and mass measurements in a single‐stage orbitrap mass spectrometer

Orbitrap technology offers a combination of different technical specifications which have not yet been achieved by other high-resolution mass spectrometry instrumentation. This refers to the combination of sensitivity, dynamic range, mass accuracy, resolution and speed. The high stability of the mass axis and the general ease of use made the orbitrap instrumentation attractive for routine laboratories. However, there are circumstances where significantly deviating relative isotopic abundance (RIA) and shifting accurate masses can be observed. RIA becomes biased at low ion counts. Furthermore, two adjacent, only partially resolved near-isobaric ions are detected with a deviating RIA. The presence of a very intensive mass peak does not only induce Fourier transformation related artefacts (side-lobes) but can cause mass shifts of small adjacent near-isobaric mass peaks. These effects are not as drastic as known for Fourier transform ion cyclotron resonance instruments. Still, users trying to identify or quantify trace level compounds should be aware about such limitations in order to avoid possible pitfalls.

A New Statistical Model and a Redefinition of Isotopic Ratio

Ultra-microanalysis of isotopes is required to clarify the origin, structure, and history of extremely small samples, for example micrometeorites and aerosols. However, a decrease in the number of measured atoms reveals a serious problem in the concept of isotopic ratio. The first fundamental defect of the isotopic ratio concept is imperfect reversibility of numerator and denominator, leading to two isotopic compositions inconsistent with each other. Secondly, the probability distribution of isotopic ratio is not the same as the binominal distribution, indicating that the mean isotopic ratio is systematically higher than the true isotopic ratio. Since these demerits should be avoided in ultra-microanalysis as much as possible, isotopic abundance or relative isotopic abundance should be used rather than isotopic ratio. The concept of isotopic ratio is, however, widely used in geochemical and cosmochemical fields, making it very difficult to abandon. Therefore, the potential uncertainty of isotopic ratio and the gap between the obtained isotopic ratio and the true one deriving from the number of sampling atoms and the isotopic ratio are vigorously evaluated here. In addition, a confirmation requirement for confirming an anomalous isotopic ratio is provided. The statistical model shown in the present work clearly indicates that the major isotope must be set as the denominator of isotopic ratio, i.e. isotopic ratio must be below 1, to reduce the effects of distorted probability distribution of isotopic ratio. In the precise measurement of isotopes, the potential error and disagreement with the true value must be tested when the number of sampled atoms is extremely few. Serious problems have not been occurred in research up to the present even in the case of NanoSIMS measurements, but this is because a sufficient number of atoms was analyzed and only extremely large isotopic anomalies were investigated. The correction and evaluation methods established in this paper will be required for accurate ultra-microanalysis.

Stable Nitrogen and Carbon Isotope Ratios Indicate Traditional and Market Food Intake in an Indigenous Circumpolar Population 3

The transition of a society from traditional to market-based diets (termed the nutrition transition) has been associated with profound changes in culture and health. We are developing biomarkers to track the nutrition transition in the Yup'ik Eskimo population of Southwest Alaska based on naturally occurring variations in the relative abundances of carbon and nitrogen stable isotopes (δ15N and δ13C values). Here, we provide three pieces of evidence toward the validation of these biomarkers. First, we analyzed the δ15N and δ13C values of a comprehensive sample of Yup'ik foods. We found that δ15N values were elevated in fish and marine mammals and that δ13C values were elevated in market foods containing corn or sugar cane carbon. Second, we evaluated the associations between RBC δ15N and δ13C values and self-reported measures of traditional and market food intake (n = 230). RBC δ15N values were correlated with intake of fish and marine mammals (r = 0.52; P < 0.0001). RBC δ13C values were correlated with intake of market foods made from corn and sugar cane (r = 0.46; P < 0.0001) and total market food intake (r = 0.46; P < 0.0001). Finally, we assessed whether stable isotope ratios captured population-level patterns of traditional and market intake (n = 1003). Isotopic biomarkers of traditional and market intake were associated with age, community location, sex, and cultural identity. Self-report methods showed variations by age and cultural identity only. Thus, stable isotopes show potential as biomarkers for monitoring dietary change in indigenous circumpolar populations.

Dynamic Proteome Analysis of Cyanothece sp. ATCC 51142 under Constant Light

Understanding the dynamic nature of protein abundances provides insights into protein turnover not readily apparent from conventional, static mass spectrometry measurements. This level of data is particularly informative when surveying protein abundances in biological systems subjected to large perturbations or alterations in environment such as cyanobacteria. Our current analysis expands upon conventional proteomic approaches in cyanobacteria by measuring dynamic changes of the proteome using a (13)C(15)N-l-leucine metabolic labeling in Cyanothece ATCC51142. Metabolically labeled Cyanothece ATCC51142 cells grown under nitrogen-sufficient conditions in continuous light were monitored longitudinally for isotope incorporation over a 48 h period, revealing 414 proteins with dynamic changes in abundances. In particular, proteins involved in carbon fixation, pentose phosphate pathway, cellular protection, redox regulation, protein folding, assembly, and degradation showed higher levels of isotope incorporation, suggesting that these biochemical pathways are important for growth under continuous light. Calculation of relative isotope abundances (RIA) values allowed the measurement of actual active protein synthesis over time for different biochemical pathways under high light exposure. Overall results demonstrated the utility of "non-steady state" pulsed metabolic labeling for systems-wide dynamic quantification of the proteome in Cyanothece ATCC51142 that can also be applied to other cyanobacteria.

A Data Processing Pipeline for Mammalian Proteome Dynamics Studies Using Stable Isotope Metabolic Labeling

In a recent study, in vivo metabolic labeling using (15)N traced the rate of label incorporation among more than 1700 proteins simultaneously and enabled the determination of individual protein turnover rate constants over a dynamic range of three orders of magnitude (Price, J. C., Guan, S., Burlingame, A., Prusiner, S. B., and Ghaemmaghami, S. (2010) Analysis of proteome dynamics in the mouse brain. Proc. Natl. Acad. Sci. U. S. A. 107, 14508-14513). These studies of protein dynamics provide a deeper understanding of healthy development and well-being of complex organisms, as well as the possible causes and progression of disease. In addition to a fully labeled food source and appropriate mass spectrometry platform, an essential and enabling component of such large scale investigations is a robust data processing and analysis pipeline, which is capable of the reduction of large sets of liquid chromatography tandem MS raw data files into the desired protein turnover rate constants. The data processing pipeline described in this contribution is comprised of a suite of software modules required for the workflow that fulfills such requirements. This software platform includes established software tools such as a mass spectrometry database search engine together with several additional, novel data processing modules specifically developed for (15)N metabolic labeling. These fulfill the following functions: (1) cross-extraction of (15)N-containing ion intensities from raw data files at varying biosynthetic incorporation times, (2) computation of peptide (15)N isotopic incorporation distributions, and (3) aggregation of relative isotope abundance curves for multiple peptides into single protein curves. In addition, processing parameter optimization and noise reduction procedures were found to be necessary in the processing modules in order to reduce propagation of errors in the long chain of the processing steps of the entire workflow.

Measurements of Thermal Neutron Capture Cross Sections of Low Abundance Stable Isotopes

We measure the thermal neutron capture cross sections of low abundance stable nuclei. The experiments are done with natural foils with known relative isotopic abundances. The cross sections can be obtained by comparing the yields of ’s from the neutron captures by various isotopes. The thermal neutron capture cross sections for isotopes with high abundances are already well measured, and we can obtain the cross sections for nuclei with low abundances which are not known or measured with large errors by simply getting the ratios of gamma yields. For the cross sections of the 152 Gd(n; ) reaction at thermal neutron energy, there are only two old measurements, 1100 100 barn and 735 10 barn, and there is a big discrepancy in the two values. We have measured the neutron capture cross section of the 152 Gd with a natural gadolinium foil. The cross section was obtained by comparing the yields of ’s from the neutron captures by 158 Gd. We have obtained a new cross section value of 614.7 57.8 barn for the 152 Gd(n; ) reaction.

Exploring the limits of robust detection of incorporation of 13C by mass spectrometry in protein-based stable isotope probing (protein-SIP)

One of the features of protein-based stable isotope probing is the parallel identification of differentially labeled peptide forms and the accurate calculation of their relative isotope abundances. The level of incorporation is informative of the metabolic activity of the species that synthesized the said protein and peptide. To model the carbon flux in a microbial community, an accurate assessment is crucial. Since the initial processes in carbon consumption are one of the most interesting objectives in microbial ecology, the methodology to detect low amounts of incorporation was tuned, and the limits of robust detection were analyzed. For this, Pseudomonas fluorescens DSM 50090T was grown on galactose using different ratios of (12)C/(13)C galactose from 10% down to 0.1% labeled galactose. After prolonged cultivation to ensure complete labeling, protein samples were separated by one-dimensional gel electrophoresis, subsequently tryptically digested and analyzed by ultra-performance liquid chromatography (UPLC) Orbitrap tandem mass spectrometry (MS/MS) measurements. The isotopic patterns from identified peptides in the mass spectra were used to calculate the (13)C relative isotope abundance (RIA) in the respective peptides. The statistic distribution of the RIA values in dependence of the number of analyzed peptides was compared between the different ratios of unlabeled/labeled substrate. The acquired data showed that the applied method is capable of detecting a difference in (13)C incorporation of ±0.1% RIA based on at least 20 peptides. This sensitivity makes protein-stable isotope probing a valuable method for quantitative assessment of species specific metabolic activity in metaproteomics.

An high resolution FDIRC for the measurement of cosmic-ray isotopic abundances

Measurements of the relative abundance of cosmic isotopes and of the energy dependence of their fluxes may clarify our present understanding on the confinement time of charged cosmic rays in the Galaxy. Experimental studies of these propagation clocks have been carried out by balloon and space missions at energies of a few 100 MeV/amu by means of detection techniques based on multiple d E/d x sampling, coupled with a measurement of the energy released in a thick absorber. At larger energies, the isotopic separation of light nuclei (as, for instance, 9Be/ 10Be) can be achieved by combining a precise measurement of the particle’s rigidity with an high resolution determination of its velocity, via the observation of the Cherenkov effect in a radiator. In this paper, we propose the introduction – for the first time in a space experiment – of the DIRC technique (Detection of Internal Reflected Cherenkov light) for the identification of cosmic-ray isotopes. This type of detector has been successfully used in electron–positron colliders for particle identification and in particular for π–K separation. While for particles with unit charge the light yield is a limiting factor, in the case of a nucleus of charge Z the larger photostatistics (due to the Z 2 dependence of Cherenkov light emission) is the key to reach an adequate angular resolution to provide a mass discrimination for isotopes of astrophysical interest. We report on the early development phase of a DIRC prototype with a focussing scheme (FDIRC) to collect the Cherenkov light onto a detector plane instrumented with a Silicon PhotoMultiplier (SiPM) array.

Characterization of Isotopic Abundance Measurements in High Resolution FT-ICR and Orbitrap Mass Spectra for Improved Confidence of Metabolite Identification

Currently there is limited information available on the accuracy and precision of relative isotopic abundance (RIA) measurements using high-resolution direct-infusion mass spectrometry (HR DIMS), and it is unclear if this information can benefit automated peak annotation in metabolomics. Here we characterize the accuracy of RIA measurements on the Thermo LTQ FT Ultra (resolution of 100,000-750,000) and LTQ Orbitrap (R = 100,000) mass spectrometers. This first involved reoptimizing the SIM-stitching method (Southam, A. D. Anal. Chem. 2007, 79, 4595-4602) for the LTQ FT Ultra, which achieved a ca. 3-fold sensitivity increase compared to the original method while maintaining a root-mean-squared mass error of 0.16 ppm. Using this method, we show the quality of RIA measurements is highly dependent on signal-to-noise ratio (SNR), with RIA accuracy increasing with higher SNR. Furthermore, a negative offset between the theoretical and empirically calculated numbers of carbon atoms was observed for both mass spectrometers. Increasing the resolution of the LTQ FT Ultra lowered both the sensitivity and the quality of RIA measurements. Overall, although the errors in the empirically calculated number of carbons can be large (e.g., 10 carbons), we demonstrate that RIA measurements do improve automated peak annotation, increasing the number of single empirical formula assignments by >3-fold compared to using accurate mass alone.

Improving Precision in Resonance Ionization Mass Spectrometry: Influence of Laser Bandwidth in Uranium Isotope Ratio Measurements

The use of broad bandwidth lasers with automated feedback control of wavelength was applied to the measurement of (235)U/(238)U ratios by resonance ionization mass spectrometry (RIMS) to decrease laser-induced isotopic fractionation. By broadening the bandwidth of the first laser in a three-color, three-photon ionization process from a bandwidth of 1.8 GHz to about 10 GHz, the variation in sequential relative isotope abundance measurements decreased from 10% to less than 0.5%. This procedure was demonstrated for the direct interrogation of uranium oxide targets with essentially no sample preparation.

Proteome Scale Turnover Analysis in Live Animals Using Stable Isotope Metabolic Labeling

At present most quantitative proteomics investigations are focused on the analysis of protein expression differences between two or more sample specimens. With each analysis a static snapshot of a cellular state is captured with regard to protein expression. However, any information on protein turnover cannot be obtained using classic methodologies. Protein turnover, the result of protein synthesis and degradation, represents a dynamic process, which is of equal importance to understanding physiological processes. Methods employing isotopic tracers have been developed to measure protein turnover. However, applying these methods to live animals is often complicated by the fact that an assessment of precursor pool relative isotope abundance is required. Also, data analysis becomes difficult in case of low label incorporation, which results in a complex convolution of labeled and unlabeled peptide mass spectrometry signals. Here we present a protein turnover analysis method that circumvents this problem using a (15)N-labeled diet as an isotopic tracer. Mice were fed with the labeled diet for limited time periods and the resulting partially labeled proteins digested and subjected to tandem mass spectrometry. For the interpretation of the mass spectrometry data, we have developed the ProTurnyzer software that allows the determination of protein fractional synthesis rates without the need of precursor relative isotope abundance information. We present results validating ProTurnyzer with Escherichia coli protein data and apply the method to mouse brain and plasma proteomes for automated turnover studies.

Tracing and Quantifying Anthropogenic Mercury Sources in Soils of Northern France Using Isotopic Signatures

The mercury (Hg) isotopic composition was investigated in topsoils from two case studies in north of France. The Hg isotope composition was first determined in agricultural topsoils contaminated by a close by Pb-Zn smelter. The Hg isotopic composition was also measured in topsoils from an urban area in northeastern France (Metz). In both cases, no significant mass independent isotope fractionation could be found in the soils. However, the soil isotopic composition (δ(202)Hg) was enriched in the heavier isotopes as the Hg concentration increased in the soils. A linear relationship between the δ(202)Hg in soils and 1/[Hg] indicated a mixing between a contamination source and the Hg derived from the geogenic background soils. Such findings demonstrate that the contamination signature was preserved in the soils and that the deposition of anthropogenic Hg was predominant compared to reactions leading to isotope fractionation such as biotic and abiotic reduction of Hg(II) and resulting in Hg mobility or evasion from the soils. It was therefore possible, for the first time in the case of Hg, to evaluate the contribution of the contamination source relative to the background Hg source in urban topsoils using relative isotope abundances.

Roles for Stress-inducible Lambda Glutathione Transferases in Flavonoid Metabolism in Plants as Identified by Ligand Fishing

The glutathione transferases (GSTs) of plants are a superfamily of abundant enzymes whose roles in endogenous metabolism are largely unknown. For example, the lambda class of GSTs (GSTLs) have members that are selectively induced by chemical stress treatments and based on their enzyme chemistry are predicted to have roles in redox homeostasis. However, using conventional approaches these functions have yet to be determined. To address this, recombinant GSTLs from wheat and Arabidopsis were tagged with a Strep tag and after affinity-immobilization, incubated with extracts from Arabidopsis, tobacco, and wheat. Bound ligands were then recovered by solvent extraction and identified by mass spectrometry (MS). With the wheat enzyme TaGSTL1, the ligand profiles obtained with in vitro extracts from tobacco closely matched those observed after the protein had been expressed in planta, demonstrating that these associations were physiologically representative. The stress-inducible TaGSTL1 was found to selectively recognize flavonols (e.g. taxifolin; K(d) = 25 nM), with this binding being dependent upon S-glutathionylation of an active site cysteine. In the case of the wheat extracts, this selectivity in ligand recognitions lead to the detection of flavonols that had not been previously described in this cereal. Subsequent in vitro assays showed that the co-binding of flavonols, such as quercetin, to the thiolated TaGSTL1 represented an intermediate step in the reduction of the respective S-glutathionylated quinone derivatives to yield free flavonols. These results suggest a novel role for GSTLs in maintaining the flavonoid pool under stress conditions.

NEUTRON-RICH CHROMIUM ISOTOPE ANOMALIES IN SUPERNOVA NANOPARTICLES

Neutron-rich isotopes with masses near that of iron are produced in type Ia and II supernovae. Traces of such nucleosynthesis are found in primitive meteorites in the form of variations in the isotopic abundance of 54Cr, the most neutron-rich stable isotope of chromium. The hosts of these isotopic anomalies must be presolar grains that condensed in the outflows of supernovae, offering the opportunity to study the nucleosynthesis of iron-peak nuclei in ways that complement spectroscopic observations and can inform models of stellar evolution. However, despite almost two decades of extensive search, the carrier of 54Cr anomalies is still unknown, presumably because it is fine-grained and is chemically labile. Here we identify in the primitive meteorite Orgueil the carrier of 54Cr-anomalies as nanoparticles, most likely spinels that show large enrichments in 54Cr relative to solar composition (54Cr/52Cr ratio >3.6xsolar). Such large enrichments in 54Cr can only be produced in supernovae. The mineralogy of the grains supports condensation in the O/Ne-O/C zones of a type II supernova, although a type Ia origin cannot be excluded. We suggest that planetary materials incorporated different amounts of these nanoparticles, possibly due to late injection by a nearby supernova that also delivered 26Al and 60Fe to the solar system. This idea explains why the relative abundance of 54Cr and other neutron-rich isotopes vary between planets and meteorites. We anticipate that future isotopic studies of the grains identified here will shed new light on the birth of the solar system and the conditions insupernovae.

Strategy for the elucidation of elemental compositions of trace analytes based on a mass resolution of 100 000 full width at half maximum

Elemental compositions (ECs) can be elucidated by evaluating the high-resolution mass spectra of unknown or suspected unfragmented analyte ions. Classical approaches utilize the exact mass of the monoisotopic peak (M + 0) and the relative abundance of isotope peaks (M + 1 and M + 2). The availability of high-resolution instruments like the Orbitrap currently permits mass resolutions up to 100,000 full width at half maximum. This not only allows the determination of relative isotopic abundances (RIAs), but also the extraction of other diagnostic information from the spectra, such as fully resolved signals originating from (34)S isotopes and fully or partially resolved signals related to (15)N isotopes (isotopic fine structure). Fully and partially resolved peaks can be evaluated by visual inspection of the measured peak profiles. This approach is shown to be capable of correctly discarding many of the EC candidates which were proposed by commercial EC calculating algorithms. Using this intuitive strategy significantly extends the upper mass range for the successful elucidation of ECs.

Evaluation of Accurate Mass and Relative Isotopic Abundance Measurements in the LTQ-Orbitrap Mass Spectrometer for Further Metabolomics Database Building

Recently, high-resolution mass spectrometry has been largely employed for compound identification, thanks to accurate mass measurements. As additional information, relative isotope abundance (RIA) is often needed to reduce the number of candidates prior to tandem MS(n). Here, we report on the evaluation of the LTQ-Orbitrap, in terms of accurate mass and RIA measurements for building further metabolomics spectral databases. Accurate mass measurements were achieved in the ppm range, using external calibration within 24 h, and remained at <5 ppm over a one-week period. The experimental relative abundances of (M+1) isotopic ions were evaluated in different data sets. First of all, 137 solutions of commercial compounds were analyzed by flow injection analysis in both the positive and negative ion modes. It was found that the ion abundance was the main factor impacting the accuracy of RIA measurements. It was possible to define some intensity thresholds above which errors were systematically <20% of their theoretical values. The same type of results were obtained with analyses from two biological media. Otherwise, no significant effect of ion transmission between the LTQ ion trap and the Orbitrap analyzer on RIA measurement errors was found, whereas the reliability of RIA measurements was dramatically improved by reducing the mass detection window. It was also observed that the signal integration method had a significant impact on RIA measurement errors, with the most-reliable results being obtained with peak height integrations. Finally, automatic integrations using the data preprocessing software XCMS and MZmine gave results similar to those obtained by manual integration, suggesting that it is relevant to use the RIA information in automatic elemental composition determination software from metabolomic peak tables.