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PubMed1983

Labeling of soybeans with the stable isotope 70Zn for use in human metabolic studies.

Radiozinc (65Zn) was used to assess the efficiency of incorporation of the label into soybean seeds by means of a noncycling hydroponic system. It was found that 21.3 and 27.6% of the applied dose was incorporated into the mature seeds when the tracer was applied during vegetative or flowering stages of growth cycle, respectively. The latter method was adapted to labeling with the stable isotope 70Zn, and quantitative incorporation of this isotope was investigated within and among hydroponic culture pots each containing four plants. It was found that on the average 17.7% +/- 3.0% of the applied dose was recovered in the mature seeds harvested from each pot. The mass isotope enrichment ratio for 70Zn: 68Zn in these seeds was 2.93 +/- 0.38 (mean +/- SD) compared with the natural ratio of 0.0343, indicating a high degree of stable isotope enrichment. Results of soybean fractionation studies showed that the enrichment was somewhat higher in the defatted meal fraction as compared with the full-fat, the hull, and the whole seed, indicating somewhat preferential translocation of the applied stable isotope in the protein. Despite this small but observable difference, all fractions were highly enriched with 70Zn. The data from this study indicated that noncycling hydroponic culture may be effectively employed to intrinsically label soybeans with the stable isotopes of zinc for use in human feeding trials dealing with the issues of zinc availability. A 100-g portion of such labeled seeds provided 38-43 g protein and 490-620 micrograms 70Zn in various processed products for use in human feeding studies.

Humans↗
PubMed2004

Heterogeneity in stable isotope profiles predicts coexistence of populations of barn swallows Hirundo rustica differing in morphology and reproductive performance.

Population studies assume that individuals belonging to a study population are homogeneous for natal and breeding origin, although this assumption is rarely tested. We tested for heterogeneity in stable-isotope profiles (delta15N, delta13C, deltaD) of feathers grown in the African winter quarters from a Danish breeding population of adult barn swallows, Hirundo rustica. Deuterium isotope values did not provide useful information on population segregation of wintering swallows in Africa. However, both delta15N and delta13C values showed a clearly bimodal distribution with 6% belonging to one category and the remaining birds belonging to another category, resulting in this population comprising three categories of birds. Adults belonging to the two categories of delta13C isotope profiles differed weakly in morphology for several different characters. The frequency and the size of second broods differed between categories of delta13C isotope profiles. Phenotypes of nestlings from the first brood in terms of tarsus length, body mass and T-cell response differed significantly between the two delta15N isotope categories, suggesting that conditions during winter carried over to the breeding season at least as late as the first brood. Polymorphism can be maintained only if fitness is similar for birds from categories of isotope profiles. We suggest that fluctuating selection or migration-selection balance may maintain the observed polymorphism.

Analysis of Variance↗
PubMed1987

Models to interpret kinetic data in stable isotope tracer studies.

In contrast to "weightless" radioactive tracers, stable isotope tracers have nonnegligible mass and are naturally present in the system, and the measured variable is a ratio of two isotopic species. These features do not allow stable isotopic tracer data analysis using straightforward analogy with radioactive tracer approaches, even though this practice is common. In this study, we present kinetic variables, models, and measurements for the analysis and interpretation of stable isotope tracer data. Assumptions and mathematical techniques for modeling the data when perturbation is both nonnegligible and negligible are discussed. Emphasis is placed on the rich information content of the dynamic portion of a stable isotope tracer curve and on the role of compartmental and noncompartmental modeling approaches for its interpretation. A presumed and commonly used analogy between the radioactive specific activity and stable isotopic enrichment is shown to be incorrect. We show that the proper analogue of specific activity is the tracer-to-tracee molar ratio. This variable is not a directly measurable one, but a formula is derived that allows its computation from the data. A method for reconstructing the time course in blood of the concentration component due to endogenous synthesis is presented. This allows measurement of the extent of the perturbation in the case where a nonweightless tracer is used. Special attention is given to data analysis originating from a multiple tracer experiment, a configuration necessary for studying more complex systems, e.g., the kinetics of interacting substrates.

3-Hydroxybutyric Acid↗
PubMed1998

Ratio encoding combinatorial libraries with stable isotopes and their utility in pharmaceutical research.

Combinatorial libraries are an important tool for lead discovery in the pharmaceutical industry. Advances in high throughput screening coupled with combinatorial chemistry can significantly reduce the time to find lead compounds. A major difficulty in developing large combinatorial libraries is the ability to identify active compounds. This paper describes a rapid and sensitive encoding/decoding methodology that utilizes stable isotopes and mass spectrometry. The ability of mass spectrometry to precisely determine the intensity of isotopic abundances provides a unique encoding strategy employing synthetically generated ratios of stable isotopes in a compound as the code. The application of ratio encoding is demonstrated using peptoid and imidazole chemistries. Supporting data demonstrate that the incorporation of one or more stable isotopes using unique-predetermined ratios can encode chemical libraries. In addition, the presence of a unique isotopic pattern in a ligand can facilitate the pharmacokinetic analysis. Isotope incorporation into a compound and subsequently into its metabolites reliably distinguishes products from other molecules in the mass spectrum. This is illustrated by metabolic analyses of peptoid and imidazole compounds.

Amino Acid Sequence↗
PubMed1996

Isotopic constraints on the age and early differentiation of the Earth.

The Earth's age and early differentiation history are re-evaluated using updated isotopic constraints. From the most primitive terrestrial Pb isotopic compositions found at Isua Greenland, and the Pilbara of Western Australia, combined with precise geochronology of these localities, an age 4.49 +/- 0.02 Ga is obtained. This is interpreted as the mean age of core formation as U/Pb is fractionated due to sequestering of Pb into the Earth's core. The long-lived Rb-Sr isotopic system provides constraints on the time interval for the accretion of the Earth as Rb underwent significant depletion by volatile loss during accretion of the Earth or its precursor planetesimals. A primitive measured 87Sr/86Sr initial ratio of 0.700502 +/- 10 has been obtained for an early Archean (3.46 Ga) barite from the Pilbara Block of Western Australia. Using conservative models for the evolution of Rb/Sr in the early Archean mantle allows an estimate to be placed on the Earth's initial Sr ratio at approximately 4.50 Ga, of 0.69940 +/- 10. This is significantly higher than that measured for the Moon (0.69900 +/- 2) or in the achondrite, Angra dos Reis (0.69894 +/- 2) and for a Rb/Sr ratio of approximately 1/2 of chondrites corresponds to a mean age for accretion of the Earth of 4.48 + /- 0.04 Ga. The now extinct 146Sm-142Nd (T1/2(146)=103 l0(6)yrs) combined with the long-lived 147Sm-143Nd isotopic systematics can also be used to provide limits on the time of early differentiation of the Earth. High precision analyses of the oldest (3.8-3.9 Ga) Archean gneisses from Greenland (Amitsoq and Akilia gneisses), and Canada (Acasta gneiss) do not show measurable (> +/- l0ppm) variations of 142Nd, in contrast to the 33 ppm 142Nd excess reported for an Archean sample. The general lack of 142Nd variations, combined with the presence of highly positive epsilon 143 values (+4.0) at 3.9 Ga, indicates that the record of large-scale Sm/Nd fractionation events was not preserved in the early-Earth from 4.56 Ga to approximately 4.3 Ga. This is consistent with large-scale planetary re-homogenisation during ongoing accretion of the Earth. The lack of isotopic anomalies in short-lived decay systems, together with the Pb and Sr isotopic constraints is thus consistent with core formation and accretion of the Earth occurring over an approximately 100 Ma interval following the formation of meteorites at 4.56 Ga.

Earth, Planet↗
PubMed1989

Oxygen isotope effects on the ribulosebisphosphate oxygenase reaction.

The oxygen isotope effect at the substrate O2 on the oxygenase reaction of ribulose bisphosphate carboxylase/oxygenase from spinach is pH and metal dependent. The pH dependence between pH 7.4 and 8.9 is different with Mg2+ (steady decrease in this isotope effect from 1.036 to 1.030) and Mn2+ (minimum isotope effect of 1.028 at pH 8.0). Deuteration of the substrate ([3-2H]ribulose bisphosphate) has no influence on the isotope effect. The results are interpreted as a direct participation of the metal ion in the oxygen-sensitive step, i.e. carbon-oxygen bond formation and the stabilization of the intermediates. In the overall reaction oxygen addition is a major rate-limiting step, and the observed isotope effect is probably close to the intrinsic oxygen isotope effect of the reaction. The basic mechanisms for carboxylation and oxygenation of ribulose bisphosphate appear to be the same.

Carbon Isotopes↗
PubMed1996

Synthesis of isotopically labelled retinal. Structural and functional studies at the atomic level of the chromophore in visual pigments.

OBJECTIVE: To describe the strategy followed in studying the structure and function of the chromophore in visual pigments at the atomic level. METHODS: A three-step strategy was followed. First, 11-cis retinal with high enrichment (99% 13C or 99% 2H) at pre-determined positions or combinations of positions was synthesized. Second, the rhodopsins with isotope label in the chromophore were prepared combining opsin with the isotopically labelled 11-cis retinal. Third, these systems were studied with isotope-sensitive physical techniques. CONCLUSIONS: Isotope labelling together with isotope-sensitive spectroscopy is a highly promising method for obtaining information at the atomic level of biologically active systems. Strategies based on specific stable isotope labelling are likely to be applicable to many research areas.

Carbon Isotopes↗
PubMed1982

The application of stable isotopes in biomedical research.

Many of the ways in which isotopes are used in biomedical research are reviewed. The use of stable isotopes in stable isotope dilution assays, for metabolite identification and in pharmacokinetic studies, is discussed and relevant examples are given to illustrate the various points made. Isotope effects and their implications for future drug design are considered. Some of the toxicity problems associated with the use of stable isotopes are also considered. Finally, a brief subjective view of possible future advances is made.

Animals↗
PubMed2003

Stable isotope variation as a tool to trace the authenticity of beef.

Organic beef coming principally from Germany was analysed for the hydrogen, carbon, oxygen, nitrogen and sulfur stable isotopic composition to test the possibility of tracing back the geographical origin. Since there is a well-known pattern of D/H and 18O/16O in meteoric water and in ground water, there is an existing link to tissue water in the beef. By including the stable isotope ratios of the other elements of life further information is available: soils show different isotope ratios of 15N/14N and 34S/32S depending on the geological composition, cultivation and atmospheric sulfur deposition. As organic farming is mainly obliged to use only their produced fodder, that ratio is reflected in the beef as well. Different organic beef samples from various German farms have been collected and analysed over nearly two years. To check the differentiation of foreign beef, samples from Argentina and Chile were also included in the study. The analyses of meat samples indicate that it is possible to trace back the region (e.g. Argentina and Germany) by using isotopes of oxygen and hydrogen. A local geographical differentiation can be done by using the stable isotopes of nitrogen and sulfur, as was demonstrated for three farms in Colonia Bay. An optimal differentiation also depends on the quality of further information (e.g. the season, kind of cattle breeding or the declaration of the local geographical origin). Certainly authenticity of beef is not only linked with the geographical origin but can also reflect the differentiation of organic and conventional farming. The fodder of organic cattle farming consists mainly of C3 plants and the use of C4 plants is more usual in conventional cattle farming. A 13C/12C ratio above -20 per thousand appears as a limit for organic farming. Increased values have to be controlled based on their authenticity.

Animals↗
PubMed2002

Differentiation of the geographical origin of durum wheat semolina samples on the basis of isotopic composition.

The identification of the geographical origin of foods is attracting great interest from consumers and producers since it may be used as a criterion for guaranteeing quality and authenticity. Stable isotope techniques can provide useful information on the origin of food products. The natural abundance isotopic ratios of carbon, oxygen and nitrogen were determined and a comparison made between the isotopic content of samples originating from Italy and those of samples originating from other countries (Canada, Turkey, Australia). A correlation was found between the isotopic composition and geographical origin of the samples. The relationship between isotopic content and the latitude has been confirmed.

Australia↗
PubMed1996

High precision isotope ratio monitoring techniques in mass spectrometry.

Isotope ratio monitoring following on-line combustion is a new method in gas chromatography/mass spectrometry (GC/MS) that allows the ratios of abundances of stable isotopes of elements such as carbon and nitrogen to be determined for individual compounds introduced via a gas chromatograph. It is the first combustion method that allows direct measurement of isotope ratios of individual molecular components of mixtures. Unlike traditional gas isotope ratio measurements made on pure samples, the method requires only very small samples, viz. several picolitres of the vapour. This paper reviews the principles and history of gas isotope ratio monitoring, focusing on the instrumentation used in isotope ratio monitoring GC/MS, including the combustion procedures used for atomization. Standardization methods and data manipulation techniques are described, as are applications to geochemistry, biology, medicine and other areas of science.

Chromatography, Liquid↗
PubMed2000

Compound-specific isotope analysis. Application to archaeology, biomedical sciences, biosynthesis, environment, extraterrestrial chemistry, food science, forensic science, humic substances, microbiology, organic geochemistry, soil science and sport.

The isotopic composition, for example, (14)C/(12)C, (13)C/(12)C, (2)H/(1)H, (15)N/(14)N and (18)O/(16)O, of the elements of matter is heterogeneous. It is ruled by physical, chemical and biological mechanisms. Isotopes can be employed to follow the fate of mineral and organic compounds during biogeochemical transformations. The determination of the isotopic composition of organic substances occurring at trace level in very complex mixtures such as sediments, soils and blood, has been made possible during the last 20 years due to the rapid development of molecular level isotopic techniques. After a brief glance at pioneering studies revealing isotopic breakthroughs at the molecular and intramolecular levels, this paper reviews selected applications of compound-specific isotope analysis in various scientific fields.

Archaeology↗
PubMed2005

Proteome dynamics in complex organisms: using stable isotopes to monitor individual protein turnover rates.

The complete definition of changes in a proteome requires information about dynamics and specifically the rate at which the individual proteins are turned over intracellularly. Whilst this can be achieved in single-cell culture using stable isotope precursors, it is more challenging to develop methods for intact animals. In this study, we show how dietary administration of stable isotope-labelled amino acids can obtain information on the relative rates of synthesis and degradation of individual proteins in a proteome. The pattern of stable isotope-labelling in tryptic peptides can be deconstructed to yield a highly reliable measure of the isotope abundance of the precursor pool, a parameter that is often difficult to acquire. We demonstrate this approach using chickens fed a semisynthetic diet containing [(2)H(8)]valine at a calculated relative isotope abundance (RIA) of 0.5. When the labelling pattern of gel-resolved muscle proteins was analyzed, the intracellular precursor isotope abundance was 0.35, consistent with dilution of the amino acid precursor pool with unlabelled amino acids derived from degradation of pre-existing proteins. However, the RIA was stable over an extended labelling window, and permitted calculation of the rates of synthesis and degradation of individual proteins isolated by gel electrophoresis. For the first time, it is feasible to contemplate the analysis of turnover of individual proteins in intact animals.

Animals↗
PubMed2003

Oxygen isotopes in nitrate: new reference materials for 18O:17O:16O measurements and observations on nitrate-water equilibration.

Despite a rapidly growing literature on analytical methods and field applications of O isotope-ratio measurements of NO(3)(-) in environmental studies, there is evidence that the reported data may not be comparable because reference materials with widely varying delta(18)O values have not been readily available. To address this problem, we prepared large quantities of two nitrate salts with contrasting O isotopic compositions for distribution as reference materials for O isotope-ratio measurements: USGS34 (KNO(3)) with low delta(18)O and USGS35 (NaNO(3)) with high delta(18)O and 'mass-independent' delta(17)O. The procedure used to produce USGS34 involved equilibration of HNO(3) with (18)O-depleted meteoric water. Nitric acid equilibration is proposed as a simple method for producing laboratory NO(3)(-) reference materials with a range of delta(18)O values and normal (mass-dependent) (18)O:(17)O:(16)O variation. Preliminary data indicate that the equilibrium O isotope-fractionation factor (alpha) between [NO(3)(-)] and H(2)O decreases with increasing temperature from 1.0215 at 22 degrees C to 1.0131 at 100 degrees C. USGS35 was purified from the nitrate ore deposits of the Atacama Desert in Chile and has a high (17)O:(18)O ratio owing to its atmospheric origin. These new reference materials, combined with previously distributed NO(3) (-) isotopic reference materials IAEA-N3 (=IAEA-NO-3) and USGS32, can be used to calibrate local laboratory reference materials for determining offset values, scale factors, and mass-independent effects on N and O isotope-ratio measurements in a wide variety of environmental NO(3)(-) samples. Preliminary analyses yield the following results (normalized with respect to VSMOW and SLAP, with reproducibilities of +/-0.2-0.3 per thousand, 1sigma): IAEA-N3 has delta(18)O = +25.6 per thousand and delta(17)O = +13.2 per thousand; USGS32 has delta(18)O = +25.7 per thousand; USGS34 has delta(18)O = -27.9 per thousand and delta(17)O = -14.8 per thousand; and USGS35 has delta(18)O = +57.5 per thousand and delta(17)O = +51.5 per thousand.

Mass Spectrometry↗
PubMed2004

Feeding fish with diets of different ratios of C3- and C4-plant-derived ingredients: a laboratory analysis with implications for the back-calculation of diet from stable isotope data.

Back-calculation of the diet is one of the most frequent applications of stable isotope techniques in animal ecology. These calculations are often based on two assumptions: a constant trophic shift for all dietary items and a linear response of the isotopic ratios to different mixtures of two isotopically distinct feeds. In a laboratory experiment, fish (Nile tilapia, Oreochromis niloticus) were fed semi-synthetic diets prepared either from wheat or corn ingredients, or from three blended diets (25, 50, 75% wheat components). Isotopic analysis of the lipid-free and lipid fraction of the fish after the experiment revealed that the trophic shift was not constant for wheat- and corn-based diets. The isotopic response to the mixed diets was not linear, leading to a statistically significant over-estimation of the corn component in the back-calculation. Both effects are in agreement with published data on the isotopic effects of C3- and C4-plant materials in the diet.

Administration, Oral↗
PubMed2005

Using a simple high-performance liquid chromatography separation and fraction collection methodology to achieve compound-specific isotopic analysis for dissolved organic compounds.

A new application for the quantitative and isotopic analyses of dissolved inorganic and dissolved organic carbon compounds has been developed. Dissolved organic matter (DOM) in natural water samples can be separated on a high-performance liquid chromatography (HPLC) column and collected as fractions. Each discrete fraction can then be analyzed using the technique of St-Jean (Rapid Commun. Mass Spectrom. 2003; 17: 419-428) with a total inorganic carbon/total organic carbon (TIC/TOC) analyzer interfaced with a continuous-flow isotope ratio mass spectrometer. Experimental data using short-chain fatty acid standards (formic, acetic, and propionic acids) show that fraction recoveries of 100% are possible and that sample integrity is maintained. 13C-isotopic analyses of products prior to and subsequent to extraction and collection show no isotopic effects associated with the methodology, and errors are well within the accepted analytical uncertainty of the IRMS. Comparison of data from pure standards and organic-rich natural waters shows that quantitative analyses still need to be done with standards that more closely imitate the matrices of the samples, in order to acquire an appropriate calibration curve. Injections of organic-rich matrices on the HPLC column did not affect fraction recovery, nor did they create high background of partially retained organic compounds slowly released from the HPLC column, and hence 13C-isotopic results are relatively unaffected. The specific limitation on this methodology is the required use of carbon-free carrier solvents due to potential memory effects associated with the TIC/TOC analyzer. Further developments of this application could make routine compound-specific isotopic analyses (CSIA) for a wider range of organic materials possible.

Carbon Isotopes↗
PubMed2005

A method for carbon stable isotope analysis of methyl halides and chlorofluorocarbons at pptv concentrations.

A pre-concentration system has been validated for use with a gas chromatography/mass spectrometry/isotope ratio mass spectrometer (GC/MS/IRMS) to determine ambient air (13)C/(12)C ratios for methyl halides (MeCl and MeBr) and chlorofluorocarbons (CFCs). The isotopic composition of specific compounds can provide useful information on their atmospheric budgets and biogeochemistry that cannot be ascertained from abundance measurements alone. Although pre-concentration systems have been previously used with a GC/MS/IRMS for atmospheric trace gas analysis, this is the first study also to report system validation tests. Validation results indicate that the pre-concentration system and subsequent separation technologies do not significantly alter the stable isotopic ratios of the target methyl halides, CFC-12 (CCl(2)F(2)) and CFC-113 (C(2)Cl(3)F(3)). Significant, but consistent, isotopic shifts of -27.5 per thousand to -25.6 per thousand do occur within the system for CFC-11 (CCl(3)F), although the shift is correctible. The method presented has the capacity to separate these target halocarbons from more than 50 other compounds in ambient air samples. Separation allows for the determination of stable carbon isotope ratios of five of these six target trace atmospheric constituents within ambient air for large volume samples (</=10 L). Representative urban air analyses from Belfast City are also presented which give carbon isotope results similar to published values for (13)C/(12)C analysis of MeCl (-39.1 per thousand) and CFC-113 (-28.1 per thousand). However, this is the first paper reporting stable carbon isotope signatures for CFC-11 (-29.4 per thousand) and CFC-12 (-37.0 per thousand).

Air Pollutants↗
PubMed2005

Carbon isotope analyses of cellulose using two different on-line techniques (elemental analysis and high-temperature pyrolysis)--a comparison.

Even though the recent development in on-line methods for the stable isotope determination in cellulose has led to a significant increase in sample throughput and decrease in sample preparation expenditure, there still is a large potential for optimizing the analytical procedures by simultaneously measuring the isotope ratios of two or even more elements. Therefore, the main objective of this study was to answer the question whether high-temperature pyrolysis (HTP) is a suitable and reliable technique for the determination of the carbon isotopic composition of cellulose simultaneously during the well-known conventional oxygen isotope analysis. This study shows that HTP of cellulose is a technique that can produce reasonable delta(13)C values, matching the requirements of most research problems related to paleoclimatology. The reproducibility of the delta values for (13)C/(12)C is better than 0.2 per thousand. Some deficiencies of the method are related to the incomplete conversion of the organic carbon in the sample to carbon monoxide. A clear isotope effect seems to be related to the non-statistical conversion of the carbon in the cellulose to CO. The extent of this effect appears to be controlled by the relative proportion of crystallized and amorphous matter in the cellulose structure. Those deficiencies can be eliminated by using an appropriate normalization and by applying the principles of identical treatment for reference materials and samples. In general, a very good agreement is achieved for carbon isotope values determined by HTP and elemental analysis (EA).

Carbon↗