WorldWideScience

Sample records for isotope labeling methods

  1. Reductive methods for isotopic labeling of antibiotics

    International Nuclear Information System (INIS)

    Champney, W.S.

    1989-01-01

    Methods for the reductive methylation of the amino groups of eight different antibiotics using 3 HCOH or H 14 COH are presented. The reductive labeling of an additional seven antibiotics by NaB 3 H 4 is also described. The specific activity of the methyl-labeled drugs was determined by a phosphocellulose paper binding assay. Two quantitative assays for these compounds based on the reactivity of the antibiotic amino groups with fluorescamine and of the aldehyde and ketone groups with 2,4-dinitrophenylhydrazine are also presented. Data on the cellular uptake and ribosome binding of these labeled compounds are also presented

  2. [Progress in stable isotope labeled quantitative proteomics methods].

    Science.gov (United States)

    Zhou, Yuan; Shan, Yichu; Zhang, Lihua; Zhang, Yukui

    2013-06-01

    Quantitative proteomics is an important research field in post-genomics era. There are two strategies for proteome quantification: label-free methods and stable isotope labeling methods which have become the most important strategy for quantitative proteomics at present. In the past few years, a number of quantitative methods have been developed, which support the fast development in biology research. In this work, we discuss the progress in the stable isotope labeling methods for quantitative proteomics including relative and absolute quantitative proteomics, and then give our opinions on the outlook of proteome quantification methods.

  3. Simple, rapid method for the preparation of isotopically labeled formaldehyde

    Science.gov (United States)

    Hooker, Jacob Matthew [Port Jefferson, NY; Schonberger, Matthias [Mains, DE; Schieferstein, Hanno [Aabergen, DE; Fowler, Joanna S [Bellport, NY

    2011-10-04

    Isotopically labeled formaldehyde (*C.sup..sctn.H.sub.2O) is prepared from labeled methyl iodide (*C.sup..sctn.H.sub.3I) by reaction with an oxygen nucleophile having a pendant leaving group. The mild and efficient reaction conditions result in good yields of *C.sup..sctn.H.sub.2O with little or no *C isotopic dilution. The simple, efficient production of .sup.11CH.sub.2O is described. The use of the .sup.11CH.sub.2O for the formation of positron emission tomography tracer compounds is described. The reaction can be incorporated into automated equipment available to radiochemistry laboratories. The isotopically labeled formaldehyde can be used in a variety of reactions to provide radiotracer compounds for imaging studies as well as for scintillation counting and autoradiography.

  4. Stable isotopes labelled compounds

    International Nuclear Information System (INIS)

    1982-09-01

    The catalogue on stable isotopes labelled compounds offers deuterium, nitrogen-15, and multiply labelled compounds. It includes: (1) conditions of sale and delivery, (2) the application of stable isotopes, (3) technical information, (4) product specifications, and (5) the complete delivery programme

  5. Melatonin labeled with hydrogen isotopes

    International Nuclear Information System (INIS)

    Dmitrevskaya, L.I.; Smushkevich, Yu.I.; Kurkovskaya, L.N.; Ponomarenko, N.K.; Suvorov, N.N.

    1989-01-01

    A study has been made of isotope exchange between melatonin and deuterium (D 2 O) or tritium (HTO) oxide under different conditions. The ease of isotope exchange for the indole ring hydrogens of melatonin in an acidic medium decreases over the series H 4 > H 2 H 6 >> H 7 , enabling the authors to process a route for production of melatonin labeled with hydrogen isotopes at positions 4,6, and 2 of the indole ring. A method has been suggested for producing melatonin labeled with hydrogen isotopes at position 2 by desulfurization of 2-(2,4-dinitro-phenylsulfenyl)melatonin at Ni(Re) (D)

  6. A new method for the labelling of proteins with radioactive arsenic isotopes

    Energy Technology Data Exchange (ETDEWEB)

    Jennewein, M. [Institute of Nuclear Chemistry, Johannes Gutenberg University of Mainz, Fritz-Strassmann-Weg 2, 55128 Mainz (Germany); Hermanne, A. [VUB Cyclotron, University of Brussels, Laarbeeklaan 103, 1090 Brussels (Belgium); Mason, R.P. [Department of Radiology, Advanced Radiological Sciences, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas (United States); Thorpe, P.E. [Department of Pharmacology and Simmons and Hamon Cancer Centers, University of Texas Southwestern Medical Center at Dallas, Dallas, TX (United States); Roesch, F. [Institute of Nuclear Chemistry, Johannes Gutenberg University of Mainz, Fritz-Strassmann-Weg 2, 55128 Mainz (Germany)]. E-mail: frank.roesch@uni-mainz.de

    2006-12-20

    Radioarsenic labelled radiopharmaceuticals could be a valuable asset to positron emission tomography. In particular, the long half-lives of {sup 72}As (T{sub 1/2}=26h) and {sup 74}As (T{sub 1/2}=17.8d) allow to investigate slow physiological or metabolical processes, like the enrichment and distribution of monoclonal antibodies (mab) in tumour tissue. In this work, a new method for the labelling of proteins with various radioactive arsenic isotopes was developed. For this purpose, two proteins, namely a chimeric IgG{sub 3} monoclonal antibody, ch3G4, directed against anionic phospholipids, and Rituxan (Rituximab), were labelled as a proof of principle with no-carrier-added radioarsenic isotopes ({sup 74}As and {sup 77}As). The developed labelling chemistry gives high yields (>99.9%), is reliable and could easily be transferred to automated labelling systems in a clinical environment. At least for the mab used in this work, this route of radioarsenic labelling does not affect the immunoreactivity of the product. The arsenic label stays stable for up to 72h at the molecular mass of the monoclonal antibody, which is in particular relevant to follow the pharmacology and pharmacokinetics of the labelled mab for several days.

  7. Doubly labeled water method: in vivo oxygen and hydrogen isotope fractionation

    International Nuclear Information System (INIS)

    Schoeller, D.A.; Leitch, C.A.; Brown, C.

    1986-01-01

    The accuracy and precision of the doubly labeled water method for measuring energy expenditure are influenced by isotope fractionation during evaporative water loss and CO 2 excretion. To characterize in vivo isotope fractionation, we collected and isotopically analyzed physiological fluids and gases. Breath and transcutaneous water vapor were isotopically fractionated. The degree of fractionation indicated that the former was fractionated under equilibrium control at 37 0 C, and the latter was kinetically fractionated. Sweat and urine were unfractionated. By use of isotopic balance models, the fraction of water lost via fractionating routes was estimated from the isotopic abundances of body water, local drinking water, and dietary solids. Fractionated water loss averaged 23% (SD = 10%) of water turnover, which agreed with our previous estimates based on metabolic rate, but there was a systematic difference between the results based on O 2 and hydrogen. Corrections for isotopic fractionation of water lost in breath and (nonsweat) transcutaneous loss should be made when using labeled water to measure water turnover or CO 2 production

  8. Isotopically labelled pyrimidines and purines

    International Nuclear Information System (INIS)

    Balaban, A.T.; Bally, I.

    1987-01-01

    Among the three diazines, pyrimidine is by far the most important one because its derivatives uracil, thymine and cytosine are constituents of the ubiquitous deoxynucleic acids (DNA) and ribonucleic acids (RNA). Other derivatives of pyrimidine without condensed rings include barbiturates, alloxan, orotic acid and thiamine or vitamin B 1 . From the polycyclic derivatives of pyrimidine such as pteridine, alloxazine, and purine, the latter, through its derivatives adenine and guanine complete the list of bases which occur in DNA and RNA: in addition, other purine derivatives such as hypoxanthine, xanthine, theobromine, theophylline, caffeine and uric acid are important natural products with biological activity. The paper presents methods for preparing isotopically labeled pyrimidines as well as purine derivatives. For convenience, the authors describe separately carbon-labeled with radioisotopes 11 C (T 1/2 = 20.3 min) and 14 C (T 1/2 = 5736 years) or the stable isotope 13 C (natural abundance 1.1%) and then hydrogen-labeled systems with the radioisotope 3 H ≡ T (T 1/2 = 12.346 years) or with the stable isotope 2 H ≡ D (natural abundance 0.015%). We do not separate stable from radioactive isotopes because the synthetic methods are identical for the same element; however, the introduction of hydrogen isotopes into organic molecules is often performed by reactions such as isotope exchange which cannot take place in the case of carbon isotopes

  9. Pharmaceuticals labelled with stable isotopes

    International Nuclear Information System (INIS)

    Krumbiegel, P.

    1986-11-01

    The relatively new field of pharmaceuticals labelled with stable isotopes is reviewed. Scientific, juridical, and ethical questions are discussed concerning the application of these pharmaceuticals in human medicine. 13 C, 15 N, and 2 H are the stable isotopes mainly utilized in metabolic function tests. Methodical contributions are given to the application of 2 H, 13 C, and 15 N pharmaceuticals showing new aspects and different states of development in the field under discussion. (author)

  10. Isotopically labelled benzodiazepines

    International Nuclear Information System (INIS)

    Liebman, A.A.

    1987-01-01

    This paper reports on the benzodiazepines which are a class of therapeutic agents. Improvements in the analytical methodology in the areas of biochemistry and pharmacology were significant, particularly in the application of chromatographic and spectroscopic techniques. In addition, the discovery and subsequent development of tritium and carbon-14 as an analytical tool in the biological sciences were essentially post-world war II phenomena. Thus, as these new chemical entities were found to be biologically active, they could be prepared in labeled form for metabolic study, biological half-life determination (pharmacokinetics), tissue distribution study, etc. This use of tracer methodology has been liberally applied to the benzodiazepines and also more recently to the study of receptor-ligand interactions, in which tritium, carbon-11 or fluorine-18 isotopes have been used. The history of benzodiazepines as medicinal agents is indeed an interesting one; an integral part of that history is their use in just about every conceivable labeled form

  11. The method for production of high purity carrier free ortophosphoric acid labeled with isotopes Phosphorus-32 and Phosphorus-33

    International Nuclear Information System (INIS)

    Abdukayumov, M.N.; Abdusalyamov, A.N.; Chistyakov, P.G.; Yuldashev, B.S.

    2001-01-01

    Extensive application for various radioactive isotopes was found in an extremity of the 20-Th century in a science and production. Labeled compounds are used with growing effectiveness in a molecular biology, gene engineering, medicine and other areas. Phosphorus-32 and Phosphorus-33 isotopes as a different labeled compounds that are used mainly in molecular biology are produced at the Radiopreparat enterprise of the Institute of Nuclear Physics of Academy of Sciences of Uzbekistan Republic. The quality of labeled preparations is very high. The specifications for above mentioned preparations corresponds to demands most of customers in different countries. P-32 or P-33 labeled orthophosphoric acid has high radiochemical purity (more than 99 %) and specific radioactivity close to theoretical. Orthophosphoric acid prepared by the described above method has radiochemical purity about 95 % and output of the target product 99%

  12. SAIL--stereo-array isotope labeling.

    Science.gov (United States)

    Kainosho, Masatsune; Güntert, Peter

    2009-11-01

    Optimal stereospecific and regiospecific labeling of proteins with stable isotopes enhances the nuclear magnetic resonance (NMR) method for the determination of the three-dimensional protein structures in solution. Stereo-array isotope labeling (SAIL) offers sharpened lines, spectral simplification without loss of information and the ability to rapidly collect and automatically evaluate the structural restraints required to solve a high-quality solution structure for proteins up to twice as large as before. This review gives an overview of stable isotope labeling methods for NMR spectroscopy with proteins and provides an in-depth treatment of the SAIL technology.

  13. Isotopic labelling with carbon-14 and tritium

    International Nuclear Information System (INIS)

    Evans, E.A.

    1980-01-01

    In this paper general methods of isotopic labelling with 14 C and with 3 H are briefly reviewed with special attention to examples of compounds likely to be of wide interest in biological research. (author)

  14. LC/MS Method for the Determination of Stable Isotope Labeled Promethazine in Human Plasma

    Science.gov (United States)

    Zuwei, Wang; Boyd, Jason; Berens, Kurt L.; Putcha, Lakshmi

    2004-01-01

    Promethazine (PMZ) is taken by astronauts orally (PO), intramuscularly (IM) or rectally (PR) for space motion sickness. LC/MS method was developed with off-line solid phase extraction to measure plasma concentrations of PMZ given as stable isotope-labeled (SIL) formulations by the three different routes of administration simultaneously. Samples (0.5ml) were loaded on to Waters Oasis HLB co-polymer cartridges and eluted with 1.0 mL methanol. HPLC separation of the eluted sample was performed using an Agilent Zorbax SB-CN column (50 x 2.1 mm) at a flow rate of 0.2 mL/min for 6 min. Acetonitrile/ ammonium acetate (30 mM) in water (3:2, v/v), pH 5.6 plus or minus 0.1, was used as the mobile phase for separation. Concentrations of PMZ, PMZ-d4 and PMZ-d7 and chlorpromazine (internal standard) were determined using a Micromass ZMD single quadrupole mass spectrometer with Electrospray Ionization (ESI). ESI mass spectra were acquired in positive ion mode with selected ion monitoring of [M+ H]dot plus. The method is rapid, reproducible and the assay specific parameters are listed in a table. A novel, sensitive and specific method for the measurement of PMZ and SIL PMZ in human plasma is reported.

  15. Melatonin labelled by hydrogen isotopes

    International Nuclear Information System (INIS)

    Dmitrevskaya, L.I.; Smushkevich, Yu.I.; Kurkovskaya, L.N.; Ponomarenko, N.K.; Suvorov, N.N.

    1988-01-01

    Isotope exchange of melatonin with deuterium (D 2 O) and tritium (HTO) oxides under different conditions is studied. Simplicity of isotope exchange of hydrogens of the indole ring of melatonin in the acidic medium decreases in series H 4 >H 2 >H 6 >>H 7 , that permits to suggest the way of melatonin preparation labelled by hydrogen isotopes in positions 4,6 and 2 of the indole ring. The way of melatonin preparation labelled by hydrogen isotopes in position 2 according to the reaction of desulfation 2-(2,4-dinitrophenylsulphenyl) melatonin at catalyst Ni(Re)(D) is suggested

  16. Testing isotopic labeling with [¹³C₆]glucose as a method of advanced glycation sites identification.

    Science.gov (United States)

    Kielmas, Martyna; Kijewska, Monika; Stefanowicz, Piotr; Szewczuk, Zbigniew

    2012-12-01

    The Maillard reaction occurring between reducing sugars and reactive amino groups of biomolecules leads to the formation of a heterogeneous mixture of compounds: early, intermediate, and advanced glycation end products (AGEs). These compounds could be markers of certain diseases and of the premature aging process. Detection of Amadori products can be performed by various methods, including MS/MS techniques and affinity chromatography on immobilized boronic acid. However, the diversity of the structures of AGEs makes detection of these compounds more difficult. The aim of this study was to test a new method of AGE identification based on isotope (13)C labeling. The model protein (hen egg lysozyme) was modified with an equimolar mixture of [(12)C(6)]glucose and [(13)C(6)]glucose and then subjected to reduction of the disulfide bridges followed by tryptic hydrolysis. The digest obtained was analyzed by LC-MS. The glycation products were identified on the basis of characteristic isotopic patterns resulting from the use of isotopically labeled glucose. This method allowed identification of 38 early Maillard reaction products and five different structures of the end glycation products. This isotopic labeling technique combined with LC-MS is a sensitive method for identification of advanced glycation end products even if their chemical structure is unknown. Copyright © 2012 Elsevier Inc. All rights reserved.

  17. A novel dual-isotope labelling method for distinguishing between soil sources of N2O

    NARCIS (Netherlands)

    Wrage, N.; Groenigen, van J.W.; Oenema, O.; Baggs, E.M.

    2005-01-01

    We present a novel O-18-N-15-enrichment method for the distinction between nitrous oxide (N2O) from nitrification, nitrifier denitrification and denitrification based on a method with single- and double-N-15-labelled ammonium nitrate. We added a new treatment with O-18-labelled water to quantify N2O

  18. Comparison of a Label-Free Quantitative Proteomic Method Based on Peptide Ion Current Area to the Isotope Coded Affinity Tag Method

    Directory of Open Access Journals (Sweden)

    Young Ah Goo

    2008-01-01

    Full Text Available Recently, several research groups have published methods for the determination of proteomic expression profiling by mass spectrometry without the use of exogenously added stable isotopes or stable isotope dilution theory. These so-called label-free, methods have the advantage of allowing data on each sample to be acquired independently from all other samples to which they can later be compared in silico for the purpose of measuring changes in protein expression between various biological states. We developed label free software based on direct measurement of peptide ion current area (PICA and compared it to two other methods, a simpler label free method known as spectral counting and the isotope coded affinity tag (ICAT method. Data analysis by these methods of a standard mixture containing proteins of known, but varying, concentrations showed that they performed similarly with a mean squared error of 0.09. Additionally, complex bacterial protein mixtures spiked with known concentrations of standard proteins were analyzed using the PICA label-free method. These results indicated that the PICA method detected all levels of standard spiked proteins at the 90% confidence level in this complex biological sample. This finding confirms that label-free methods, based on direct measurement of the area under a single ion current trace, performed as well as the standard ICAT method. Given the fact that the label-free methods provide ease in experimental design well beyond pair-wise comparison, label-free methods such as our PICA method are well suited for proteomic expression profiling of large numbers of samples as is needed in clinical analysis.

  19. Assessing of distribution, mobility and bioavailability of exogenous Pb in agricultural soils using isotopic labeling method coupled with BCR approach

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Zhi-Yong, E-mail: zhyhuang@jmu.edu.cn [College of Bioengineering, Jimei University, Xiamen 361021 (China); Xie, Hong [College of Bioengineering, Jimei University, Xiamen 361021 (China); Shandong Vocational Animal Science and Veterinary College, Weifang 261061 (China); Cao, Ying-Lan [College of Bioengineering, Jimei University, Xiamen 361021 (China); Cai, Chao [Key Laboratory of Urban Environment and Health, Institute of Urban Environment, Chinese Academy of Sciences, Xiamen 361021 (China); Zhang, Zhi [College of Bioengineering, Jimei University, Xiamen 361021 (China)

    2014-02-15

    Highlights: • Large amounts of exogenous Pb were found to distribute in reducible fractions. • Very few of exogenous Pb were found to distribute in acid-extractable fractions. • More than 60% of exogenous Pb in rhizosphere soils lost after planting. • Isotopic labeling method and SEP enable to explore Pb bioavailability in soil. -- Abstract: The contamination of Pb in agricultural soils is one of the most important ecological problems, which potentially results in serious health risk on human health through food chain. Hence, the fate of exogenous Pb contaminated in agricultural soils is needed to be deeply explored. By spiking soils with the stable enriched isotopes of {sup 206}Pb, the contamination of exogenous Pb{sup 2+} ions in three agricultural soils sampled from the estuary areas of Jiulong River, China was simulated in the present study, and the distribution, mobility and bioavailability of exogenous Pb in the soils were investigated using the isotopic labeling method coupled with a four-stage BCR (European Community Bureau of Reference) sequential extraction procedure. Results showed that about 60–85% of exogenous Pb was found to distribute in reducible fractions, while the exogenous Pb in acid-extractable fractions was less than 1.0%. After planting, the amounts of exogenous Pb presenting in acid-extractable, reducible and oxidizable fractions in rhizospheric soils decreased by 60–66%, in which partial exogenous Pb was assimilated by plants while most of the metal might transfer downward due to daily watering and applying fertilizer. The results show that the isotopic labeling technique coupled with sequential extraction procedures enables us to explore the distribution, mobility and bioavailability of exogenous Pb contaminated in soils, which may be useful for the further soil remediation.

  20. Assessing of distribution, mobility and bioavailability of exogenous Pb in agricultural soils using isotopic labeling method coupled with BCR approach

    International Nuclear Information System (INIS)

    Huang, Zhi-Yong; Xie, Hong; Cao, Ying-Lan; Cai, Chao; Zhang, Zhi

    2014-01-01

    Highlights: • Large amounts of exogenous Pb were found to distribute in reducible fractions. • Very few of exogenous Pb were found to distribute in acid-extractable fractions. • More than 60% of exogenous Pb in rhizosphere soils lost after planting. • Isotopic labeling method and SEP enable to explore Pb bioavailability in soil. -- Abstract: The contamination of Pb in agricultural soils is one of the most important ecological problems, which potentially results in serious health risk on human health through food chain. Hence, the fate of exogenous Pb contaminated in agricultural soils is needed to be deeply explored. By spiking soils with the stable enriched isotopes of 206 Pb, the contamination of exogenous Pb 2+ ions in three agricultural soils sampled from the estuary areas of Jiulong River, China was simulated in the present study, and the distribution, mobility and bioavailability of exogenous Pb in the soils were investigated using the isotopic labeling method coupled with a four-stage BCR (European Community Bureau of Reference) sequential extraction procedure. Results showed that about 60–85% of exogenous Pb was found to distribute in reducible fractions, while the exogenous Pb in acid-extractable fractions was less than 1.0%. After planting, the amounts of exogenous Pb presenting in acid-extractable, reducible and oxidizable fractions in rhizospheric soils decreased by 60–66%, in which partial exogenous Pb was assimilated by plants while most of the metal might transfer downward due to daily watering and applying fertilizer. The results show that the isotopic labeling technique coupled with sequential extraction procedures enables us to explore the distribution, mobility and bioavailability of exogenous Pb contaminated in soils, which may be useful for the further soil remediation

  1. Assessing of distribution, mobility and bioavailability of exogenous Pb in agricultural soils using isotopic labeling method coupled with BCR approach.

    Science.gov (United States)

    Huang, Zhi-Yong; Xie, Hong; Cao, Ying-Lan; Cai, Chao; Zhang, Zhi

    2014-02-15

    The contamination of Pb in agricultural soils is one of the most important ecological problems, which potentially results in serious health risk on human health through food chain. Hence, the fate of exogenous Pb contaminated in agricultural soils is needed to be deeply explored. By spiking soils with the stable enriched isotopes of (206)Pb, the contamination of exogenous Pb(2+) ions in three agricultural soils sampled from the estuary areas of Jiulong River, China was simulated in the present study, and the distribution, mobility and bioavailability of exogenous Pb in the soils were investigated using the isotopic labeling method coupled with a four-stage BCR (European Community Bureau of Reference) sequential extraction procedure. Results showed that about 60-85% of exogenous Pb was found to distribute in reducible fractions, while the exogenous Pb in acid-extractable fractions was less than 1.0%. After planting, the amounts of exogenous Pb presenting in acid-extractable, reducible and oxidizable fractions in rhizospheric soils decreased by 60-66%, in which partial exogenous Pb was assimilated by plants while most of the metal might transfer downward due to daily watering and applying fertilizer. The results show that the isotopic labeling technique coupled with sequential extraction procedures enables us to explore the distribution, mobility and bioavailability of exogenous Pb contaminated in soils, which may be useful for the further soil remediation. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. Gluconeogenesis from labeled carbon: estimating isotope dilution

    International Nuclear Information System (INIS)

    Kelleher, J.K.

    1986-01-01

    To estimate the rate of gluconeogenesis from steady-state incorporation of labeled 3-carbon precursors into glucose, isotope dilution must be considered so that the rate of labeling of glucose can be quantitatively converted to the rate of gluconeogenesis. An expression for the value of this isotope dilution can be derived using mathematical techniques and a model of the tricarboxylic acid (TCA) cycle. The present investigation employs a more complex model than that used in previous studies. This model includes the following pathways that may affect the correction for isotope dilution: 1) flux of 3-carbon precursor to the oxaloacetate pool via acetyl-CoA and the TCA cycle; 2) flux of 4- or 5-carbon compounds into the TCA cycle; 3) reversible flux between oxaloacetate (OAA) and pyruvate and between OAA and fumarate; 4) incomplete equilibrium between OAA pools; and 5) isotope dilution of 3-carbon tracers between the experimentally measured pool and the precursor for the TCA-cycle OAA pool. Experimental tests are outlined which investigators can use to determine whether these pathways are significant in a specific steady-state system. The study indicated that flux through these five pathways can significantly affect the correction for isotope dilution. To correct for the effects of these pathways an alternative method for calculating isotope dilution is proposed using citrate to relate the specific activities of acetyl-CoA and OAA

  3. Direct methods and residue type specific isotope labeling in NMR structure determination and model-driven sequential assignment

    International Nuclear Information System (INIS)

    Schedlbauer, Andreas; Auer, Renate; Ledolter, Karin; Tollinger, Martin; Kloiber, Karin; Lichtenecker, Roman; Ruedisser, Simon; Hommel, Ulrich; Schmid, Walther; Konrat, Robert; Kontaxis, Georg

    2008-01-01

    Direct methods in NMR based structure determination start from an unassigned ensemble of unconnected gaseous hydrogen atoms. Under favorable conditions they can produce low resolution structures of proteins. Usually a prohibitively large number of NOEs is required, to solve a protein structure ab-initio, but even with a much smaller set of distance restraints low resolution models can be obtained which resemble a protein fold. One problem is that at such low resolution and in the absence of a force field it is impossible to distinguish the correct protein fold from its mirror image. In a hybrid approach these ambiguous models have the potential to aid in the process of sequential backbone chemical shift assignment when 13 C β and 13 C' shifts are not available for sensitivity reasons. Regardless of the overall fold they enhance the information content of the NOE spectra. These, combined with residue specific labeling and minimal triple-resonance data using 13 C α connectivity can provide almost complete sequential assignment. Strategies for residue type specific labeling with customized isotope labeling patterns are of great advantage in this context. Furthermore, this approach is to some extent error-tolerant with respect to data incompleteness, limited precision of the peak picking, and structural errors caused by misassignment of NOEs

  4. Isotopically labelled vitamin D derivatives and processes for preparing same

    International Nuclear Information System (INIS)

    Deluca, H.R.; Schnoes, H.K.; Napoli, J.L.; Fivizzani, M.A.

    1981-01-01

    This invention relates to 26,27-isotopically labelled vitamin D 3 compounds, including radiolabelled vitamin D 3 compounds of high specific activity, methods for their preparation, and intermediates obtained in their synthesis. The method involves reacting an ester of a 26,27-dinor-vitamin D-25-carboxylic acid with an isotopically labelled methyl Grignard reagent or methyl lithium reagent to obtain a 26,27-isotopically labelled compound in which at least some of the H atoms and/or C atoms are heavy isotopes. (author)

  5. Validation of the doubly labeled water method using off-axis integrated cavity output spectroscopy and isotope ratio mass spectrometry.

    Science.gov (United States)

    Melanson, Edward L; Swibas, Tracy; Kohrt, Wendy M; Catenacci, Vicki A; Creasy, Seth A; Plasqui, Guy; Wouters, Loek; Speakman, John R; Berman, Elena S F

    2018-02-01

    When the doubly labeled water (DLW) method is used to measure total daily energy expenditure (TDEE), isotope measurements are typically performed using isotope ratio mass spectrometry (IRMS). New technologies, such as off-axis integrated cavity output spectroscopy (OA-ICOS) provide comparable isotopic measurements of standard waters and human urine samples, but the accuracy of carbon dioxide production (V̇co 2 ) determined with OA-ICOS has not been demonstrated. We compared simultaneous measurement V̇co 2 obtained using whole-room indirect calorimetry (IC) with DLW-based measurements from IRMS and OA-ICOS. Seventeen subjects (10 female; 22 to 63 yr) were studied for 7 consecutive days in the IC. Subjects consumed a dose of 0.25 g H 2 18 O (98% APE) and 0.14 g 2 H 2 O (99.8% APE) per kilogram of total body water, and urine samples were obtained on days 1 and 8 to measure average daily V̇co 2 using OA-ICOS and IRMS. V̇co 2 was calculated using both the plateau and intercept methods. There were no differences in V̇co 2 measured by OA-ICOS or IRMS compared with IC when the plateau method was used. When the intercept method was used, V̇co 2 using OA-ICOS did not differ from IC, but V̇co 2 measured using IRMS was significantly lower than IC. Accuracy (~1-5%), precision (~8%), intraclass correlation coefficients ( R = 0.87-90), and root mean squared error (30-40 liters/day) of V̇co 2 measured by OA-ICOS and IRMS were similar. Both OA-ICOS and IRMS produced measurements of V̇co 2 with comparable accuracy and precision compared with IC.

  6. NMR studies of isotopically labeled RNA

    Energy Technology Data Exchange (ETDEWEB)

    Pardi, A. [Univ. of Colorado, Boulder, CO (United States)

    1994-12-01

    In summary, the ability to generate NMR quantities of {sup 15}N and {sup 13}C-labeled RNAs has led to the development of heteronuclear multi-dimensional NMR techniques for simplifying the resonance assignment and structure determination of RNAs. These methods for synthesizing isotopically labeled RNAs are only several years old, and thus there are still relatively few applications of heteronuclear multi-dimensional NMR techniques to RNA. However, given the critical role that RNAs play in cellular function, one can expect to see an increasing number of NMR structural studies of biologically active RNAs.

  7. Isotope methods in hydrology

    International Nuclear Information System (INIS)

    Moser, H.; Rauert, W.

    1980-01-01

    Of the investigation methods used in hydrology, tracer methods hold a special place as they are the only ones which give direct insight into the movement and distribution processes taking place in surface and ground waters. Besides the labelling of water with salts and dyes, as in the past, in recent years the use of isotopes in hydrology, in water research and use, in ground-water protection and in hydraulic engineering has increased. This by no means replaces proven methods of hydrological investigation but tends rather to complement and expand them through inter-disciplinary cooperation. The book offers a general introduction to the application of various isotope methods to specific hydrogeological and hydrological problems. The idea is to place the hydrogeologist and the hydrologist in the position to recognize which isotope method will help him solve his particular problem or indeed, make a solution possible at all. He should also be able to recognize what the prerequisites are and what work and expenditure the use of such methods involves. May the book contribute to promoting cooperation between hydrogeologists, hydrologists, hydraulic engineers and isotope specialists, and thus supplement proven methods of investigation in hydrological research and water utilization and protection wherever the use of isotope methods proves to be of advantage. (orig./HP) [de

  8. A facile method for steroid labeling by heavy isotopes of hydrogen

    Czech Academy of Sciences Publication Activity Database

    Marek, Aleš; Klepetářová, Blanka; Elbert, Tomáš

    2015-01-01

    Roč. 71, č. 29 (2015), s. 4874-4882 ISSN 0040-4020 R&D Projects: GA AV ČR IAA400550801 Institutional support: RVO:61388963 Keywords : brassinosteroids * alpha-hydroxy ketones * reductive dehalogenation * tritium * labeled compounds Subject RIV: CC - Organic Chemistry Impact factor: 2.645, year: 2015

  9. ISOTOPE METHODS IN HOMOGENEOUS CATALYSIS.

    Energy Technology Data Exchange (ETDEWEB)

    BULLOCK,R.M.; BENDER,B.R.

    2000-12-01

    The use of isotope labels has had a fundamentally important role in the determination of mechanisms of homogeneously catalyzed reactions. Mechanistic data is valuable since it can assist in the design and rational improvement of homogeneous catalysts. There are several ways to use isotopes in mechanistic chemistry. Isotopes can be introduced into controlled experiments and followed where they go or don't go; in this way, Libby, Calvin, Taube and others used isotopes to elucidate mechanistic pathways for very different, yet important chemistries. Another important isotope method is the study of kinetic isotope effects (KIEs) and equilibrium isotope effect (EIEs). Here the mere observation of where a label winds up is no longer enough - what matters is how much slower (or faster) a labeled molecule reacts than the unlabeled material. The most careti studies essentially involve the measurement of isotope fractionation between a reference ground state and the transition state. Thus kinetic isotope effects provide unique data unavailable from other methods, since information about the transition state of a reaction is obtained. Because getting an experimental glimpse of transition states is really tantamount to understanding catalysis, kinetic isotope effects are very powerful.

  10. Stable isotope dimethyl labelling for quantitative proteomics and beyond

    Science.gov (United States)

    Hsu, Jue-Liang; Chen, Shu-Hui

    2016-01-01

    Stable-isotope reductive dimethylation, a cost-effective, simple, robust, reliable and easy-to- multiplex labelling method, is widely applied to quantitative proteomics using liquid chromatography-mass spectrometry. This review focuses on biological applications of stable-isotope dimethyl labelling for a large-scale comparative analysis of protein expression and post-translational modifications based on its unique properties of the labelling chemistry. Some other applications of the labelling method for sample preparation and mass spectrometry-based protein identification and characterization are also summarized. This article is part of the themed issue ‘Quantitative mass spectrometry’. PMID:27644970

  11. Synthesis of 15N isotope labeled alanine

    International Nuclear Information System (INIS)

    Oliveira, Claudineia R. de; Bendassolli, Jose Albertino; Sant'Ana, Carlos Roberto; Tagliassachi, Romulo Barbieri; Maximo, Everaldo; Prestes, Clelber Vieira

    2005-01-01

    The application of light chemical elements and their stable isotopes in biological studies have been increased over the last years. The use of 15 N labeled amino acids is an important tool for elucidation of peptides structures. This paper describe a method for the synthesis of 15 N isotope labeled alanine at lower costs than international ones, as well as the details of the recovery system of the nitrogen residues. In the present work an amination of α-haloacids, with the bromopropionic carboxylic acid and labeled aqua ammonia ( 15 NH 3 aq) was carried out. In order to avoid eventually losses of 15 NH 3 , special cares were adopted, since the production cost is high. Although the acquisition cost of the 13 N (radioactive) labeled compounds is lower, the obtained stable tracer will allow the accomplishment of important studies of the nitrogen cycling in living things, less occupational and environment hazards, and the time limitation problems in field studies. The tests took place in triplicates with NH 3 (aq) being employed. With the establishment of the system for 15 NH 3 recovery, an average of 94 % of the ammonia employed in the synthesis process was recovered. The purity of the amino acid was state determined by TLC (Thin Layer Chromatography) and HPLC (High-Performance Liquid Chromatography) with a fluorescence detector. The Rf and the retention time of the synthesized sample were similar the sigma standard. Finally, regarding the established conditions, it was possible to obtain the alanine with a production cost about 40 % lower than the international price. (author)

  12. Protein labelling with stable isotopes: strategies

    International Nuclear Information System (INIS)

    Lirsac, P.N.; Gilles, N.; Jamin, N.; Toma, F.; Gabrielsen, O.; Boulain, J.C.; Menez, A.

    1994-01-01

    A protein labelling technique with stable isotopes has been developed at the CEA: a labelled complete medium has been developed, performing as well as the Luria medium, but differing from it because it contains not only free aminated acids and peptides, but also sugars (96% of D-glucopyrannose) and labelled nucleosides. These precursors are produced from a labelled photosynthetic micro-organisms biomass, obtained with micro-algae having incorporated carbon 13, nitrogen 15 and deuterium during their culture. Labelling costs are reduced. 1 fig., 1 tab., 3 refs

  13. An isotope approach based on C-13 pulse-chase labelling vs. the root trenching method to separate heterotrophic and autotrophic respiration in cultivated peatlands

    Energy Technology Data Exchange (ETDEWEB)

    Biasi, C.; Pitkamaki, A. S.; Tavi, N. M.; Koponen, H. T.; Martikainen, P. J. [Univ.of Eastern Finland, Kuopio (Finland). Dept. of Environmental Science], e-mail: christina.biasi@uef.fi

    2012-11-01

    We tested an isotope method based on C-13 pulse-chase labelling for determining the fractional contribution of soil microbial respiration to overall soil respiration in an organic soil (cutaway peatland, eastern Finland), cultivated with the bioenergy crop, reed canary grass. The plants were exposed to CO{sub 2}-13 for five hours and the label was thereafter determined in CO{sub 2} derived from the soil-root system. A two-pool isotope mixing model was used to separate sources of respiration. The isotopic approach showed that a minimum of 50% of the total CO{sub 2} originated from soil-microbial respiration. Even though the method uses undisturbed soil-plant systems, it has limitations concerning the experimental determination of the true isotopic signal of all components contributing to autotrophic respiration. A trenching experiment which was comparatively conducted resulted in a 71% fractional contribution of soil-microbial respiration. This value was likely overestimated. Further studies are needed to evaluate critically the output from these two partitioning approaches. (orig.)

  14. Preparation of radioactive labelled compounds. Pt. 2. 82Br labelled organic bromine compounds by isotopic exchange

    International Nuclear Information System (INIS)

    Otto, R.

    1988-05-01

    Studies on isotopic exchange between organic bromine compounds and 82 Br labelled dioxane dibromide in the presence of AlCl 3 are described. The results obtained enable to develop a simple and quick preparation method for the labelling with 82 Br [fr

  15. Method for separating isotopes

    International Nuclear Information System (INIS)

    Jepson, B.E.

    1976-01-01

    The invention comprises a method for separating different isotopes of elements from each other by contacting a feed solution containing the different isotopes with a macrocyclic polyether to preferentially form a macrocyclic polyether complex with the lighter of the different isotopes. The macrocyclic polyether complex is then separated from the lighter isotope depleted feed solution. A chemical separation of isotopes is carried out in which a constant refluxing system permits a continuous countercurrent liquid-liquid extraction. (LL)

  16. Synthesis of isotopically labelled salicylates

    International Nuclear Information System (INIS)

    Hawkins, D.R.; Pryor, R.W.

    1981-01-01

    [ 13 C-carboxyl]Salicylic acid has been prepared by carbonation of 2-benzyloxybromobenzene followed by reductive debenzylation. Deuterium and tritium labelled salicylic acid and 2 H 2 / 13 C-salicylic acid were prepared by reduction of the 3,5-dibromo derivatives using Raney Ni-Al. Deuterium labelled salicylic acid containing up to four deuterium atoms was prepared by catalytic exchange with Raney Ni-Al in 5% NaOD/D 2 O. (author)

  17. Synthesis of isotopically labeled ketamine

    OpenAIRE

    Stuchlíková, Lucie

    2011-01-01

    In this work were synthesized ketamine isotopomers. Ketamine is used in human medicine and veterinary sectors. It has very broad spectrum of pharmacological effects: anesthetic, analgesic, hallucinogenic, bronchodilator, cardiovascular and antidepressive, which is currently in the research. At first was synthesized precursor of ketamine, N- desmethylketamine which was subsequently labeled the deuterium, tritium and carbon- 14. For the determination of purity and identity mass spectrometry and...

  18. Advances in stable isotope assisted labeling strategies with information science.

    Science.gov (United States)

    Kigawa, Takanori

    2017-08-15

    Stable-isotope (SI) labeling of proteins is an essential technique to investigate their structures, interactions or dynamics by nuclear magnetic resonance (NMR) spectroscopy. The assignment of the main-chain signals, which is the fundamental first step in these analyses, is usually achieved by a sequential assignment method based on triple resonance experiments. Independently of the triple resonance experiment-based sequential assignment, amino acid-selective SI labeling is beneficial for discriminating the amino acid type of each signal; therefore, it is especially useful for the signal assignment of difficult targets. Various combinatorial selective labeling schemes have been developed as more sophisticated labeling strategies. In these strategies, amino acids are represented by combinations of SI labeled samples, rather than simply assigning one amino acid to one SI labeled sample as in the case of conventional amino acid-selective labeling. These strategies have proven to be useful for NMR analyses of difficult proteins, such as those in large complex systems, in living cells, attached or integrated into membranes, or with poor solubility. In this review, recent advances in stable isotope assisted labeling strategies will be discussed. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Innovative method for carbon dioxide determination in human postmortem cardiac gas samples using headspace-gas chromatography–mass spectrometry and stable labeled isotope as internal standard

    International Nuclear Information System (INIS)

    Varlet, V.; Smith, F.; Froidmont, S. de; Dominguez, A.; Rinaldi, A.; Augsburger, M.; Mangin, P.; Grabherr, S.

    2013-01-01

    Graphical abstract: -- Highlights: •We developed a method for CO 2 analysis in cardiac samples and quantification by 13 CO 2 . •This method was fully validated by accuracy profile. •We have applied this method to perform CO 2 precise quantification for forensic applications. •Context of the death could be documented following CO 2 concentrations. -- Abstract: A novel approach to measure carbon dioxide (CO 2 ) in gaseous samples, based on a precise and accurate quantification by 13 CO 2 internal standard generated in situ is presented. The main goal of this study was to provide an innovative headspace-gas chromatography–mass spectrometry (HS-GC–MS) method applicable in the routine determination of CO 2 . The main drawback of the GC methods discussed in the literature for CO 2 measurement is the lack of a specific internal standard necessary to perform quantification. CO 2 measurement is still quantified by external calibration without taking into account analytical problems which can often occur considering gaseous samples. To avoid the manipulation of a stable isotope-labeled gas, we have chosen to generate in situ an internal labeled standard gas ( 13 CO 2 ) on the basis of the stoichiometric formation of CO 2 by the reaction of hydrochloric acid (HCl) with sodium hydrogen carbonate (NaH 13 CO 3 ). This method allows a precise measurement of CO 2 concentration and was validated on various human postmortem gas samples in order to study its efficiency

  20. An isotope-labeled chemical derivatization method for the quantitation of short-chain fatty acids in human feces by liquid chromatography–tandem mass spectrometry

    International Nuclear Information System (INIS)

    Han, Jun; Lin, Karen; Sequeira, Carita; Borchers, Christoph H.

    2015-01-01

    Highlights: • 3-Nitrophenylhydrazine was used to derivatize short-chain fatty acids (SCFAs) for LC-MS/MS. • 13 C 6 analogues were produced for use as isotope-labeled internal standards. • Isotope-labeled standards compensate for ESI matrix effects in LC-MS/MS. • Femtomolar sensitivities and 93–108% quantitation accuracy were achieved for human fecal SCFAs. - Abstract: Short-chain fatty acids (SCFAs) are produced by anaerobic gut microbiota in the large bowel. Qualitative and quantitative measurements of SCFAs in the intestinal tract and the fecal samples are important to understand the complex interplay between diet, gut microbiota and host metabolism homeostasis. To develop a new LC-MS/MS method for sensitive and reliable analysis of SCFAs in human fecal samples, 3-nitrophenylhydrazine (3NPH) was employed for pre-analytical derivatization to convert ten C 2 –C 6 SCFAs to their 3-nitrophenylhydrazones under a single set of optimized reaction conditions and without the need of reaction quenching. The derivatives showed excellent in-solution chemical stability. They were separated on a reversed-phase C 18 column and quantitated by negative-ion electrospray ionization – multiple-reaction monitoring (MRM)/MS. To achieve accurate quantitation, the stable isotope-labeled versions of the derivatives were synthesized in a single reaction vessel from 13 C 6 -3NPH, and were used as internal standard to compensate for the matrix effects in ESI. Method validation showed on-column limits of detection and quantitation over the range from low to high femtomoles for the ten SCFAs, and the intra-day and inter-day precision for determination of nine of the ten SCFAs in human fecal samples was ≤8.8% (n = 6). The quantitation accuracy ranged from 93.1% to 108.4% (CVs ≤ 4.6%, n = 6). This method was used to determine the SCFA concentrations and compositions in six human fecal samples. One of the six samples, which was collected from a clinically diagnosed type 2

  1. An isotope-labeled chemical derivatization method for the quantitation of short-chain fatty acids in human feces by liquid chromatography–tandem mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Han, Jun; Lin, Karen; Sequeira, Carita [University of Victoria – Genome BC Proteomics Centre, University of Victoria, Vancouver Island Technology Park, 3101–4464 Markham Street, Victoria, BC V8Z 7X8 (Canada); Borchers, Christoph H., E-mail: christoph@proteincentre.com [University of Victoria – Genome BC Proteomics Centre, University of Victoria, Vancouver Island Technology Park, 3101–4464 Markham Street, Victoria, BC V8Z 7X8 (Canada); Department of Biochemistry and Microbiology, University of Victoria, Petch Building Room 207, 3800 Finnerty Road, Victoria, BC V8P 5C2 (Canada)

    2015-01-07

    Highlights: • 3-Nitrophenylhydrazine was used to derivatize short-chain fatty acids (SCFAs) for LC-MS/MS. • {sup 13}C{sub 6} analogues were produced for use as isotope-labeled internal standards. • Isotope-labeled standards compensate for ESI matrix effects in LC-MS/MS. • Femtomolar sensitivities and 93–108% quantitation accuracy were achieved for human fecal SCFAs. - Abstract: Short-chain fatty acids (SCFAs) are produced by anaerobic gut microbiota in the large bowel. Qualitative and quantitative measurements of SCFAs in the intestinal tract and the fecal samples are important to understand the complex interplay between diet, gut microbiota and host metabolism homeostasis. To develop a new LC-MS/MS method for sensitive and reliable analysis of SCFAs in human fecal samples, 3-nitrophenylhydrazine (3NPH) was employed for pre-analytical derivatization to convert ten C{sub 2}–C{sub 6} SCFAs to their 3-nitrophenylhydrazones under a single set of optimized reaction conditions and without the need of reaction quenching. The derivatives showed excellent in-solution chemical stability. They were separated on a reversed-phase C{sub 18} column and quantitated by negative-ion electrospray ionization – multiple-reaction monitoring (MRM)/MS. To achieve accurate quantitation, the stable isotope-labeled versions of the derivatives were synthesized in a single reaction vessel from {sup 13}C{sub 6}-3NPH, and were used as internal standard to compensate for the matrix effects in ESI. Method validation showed on-column limits of detection and quantitation over the range from low to high femtomoles for the ten SCFAs, and the intra-day and inter-day precision for determination of nine of the ten SCFAs in human fecal samples was ≤8.8% (n = 6). The quantitation accuracy ranged from 93.1% to 108.4% (CVs ≤ 4.6%, n = 6). This method was used to determine the SCFA concentrations and compositions in six human fecal samples. One of the six samples, which was collected from a

  2. Isotope labeling strategies for NMR studies of RNA

    International Nuclear Information System (INIS)

    Lu, Kun; Miyazaki, Yasuyuki; Summers, Michael F.

    2010-01-01

    The known biological functions of RNA have expanded in recent years and now include gene regulation, maintenance of sub-cellular structure, and catalysis, in addition to propagation of genetic information. As for proteins, RNA function is tightly correlated with structure. Unlike proteins, structural information for larger, biologically functional RNAs is relatively limited. NMR signal degeneracy, relaxation problems, and a paucity of long-range 1 H- 1 H dipolar contacts have limited the utility of traditional NMR approaches. Selective isotope labeling, including nucleotide-specific and segmental labeling strategies, may provide the best opportunities for obtaining structural information by NMR. Here we review methods that have been developed for preparing and purifying isotopically labeled RNAs, as well as NMR strategies that have been employed for signal assignment and structure determination.

  3. Development of an isotope labeling ultra-high performance liquid chromatography mass spectrometric method for quantification of acylglycines in human urine

    Energy Technology Data Exchange (ETDEWEB)

    Stanislaus, Avalyn; Guo, Kevin [Department of Chemistry, University of Alberta, Edmonton, Alberta (Canada); Li Liang, E-mail: Liang.Li@ualberta.ca [Department of Chemistry, University of Alberta, Edmonton, Alberta (Canada)

    2012-10-31

    Graphical abstract: - Abstract: Acylglycines play a crucial regulatory and detoxification role in the accumulation of the corresponding acyl CoA esters and are an important class of metabolites in the diagnoses of inborn errors of metabolism. Sensitive quantification of a large number of acylglycines not only improves diagnosis but also enables the discovery of potential new biomarkers of diseases. We report an ultra-high performance liquid chromatography tandem mass spectrometry (UPLC-MS) method for quantifying acylglycines in human urine with high sensitivity. This method is based on the use of a newly developed isotope labeling reagent, p-dimethylaminophenacyl (DmPA) bromide, to label acylglycines to improve detection sensitivity. Eighteen acylglycines, namely acetylglycine, propionylglycine, isobutyrylglycine, butyrylglycine, 4-hydroxyphenylacetylglycine, 2-furoylglycine, tiglylglycine, 2-methybutyrylglycine, 3-methylcrotonylglycine, isovalerylglycine, valerylglycine, hexanoylglycine, phenylacetylglycine, phenylpropionylglycine, glutarylglycine, heptanoylglycine, octanoylglycine and suberylglycine, were measured. This method uses calibration standards prepared in surrogate matrix (un-derivatized urine) and stable-isotope labeled analytes as the internal standards. The analysis was carried out in the positive ion detection mode using multiple reaction monitoring (MRM) survey scans. The calibration curves were validated over the range of 1.0-500 nM. The method achieved a lower limit of quantitation (LLOQ) of 1-5 nM for all analytes, as measured by the standard derivations associated with calibration curves and confirmed in surrogate matrix; the signal-to-noise ratio at LLOQ ranged from 12.50 to 156.70. Both accuracy (% RE or relative error) and precision (% CV) were <15%. Matrix effects were minimized using the surrogate matrix. All eighteen analytes were stable in urine for at least 5 h at room temperature, autosampler (4 Degree-Sign C) for 24 h, 7 weeks at -20

  4. Syntheses with isotopically labelled carbon. Methyl iodide, formaldehyde and cyanide

    International Nuclear Information System (INIS)

    Finn, R.D.; Boothe, T.E.; Vora, M.M.; Hildner, J.C.; Emran, A.M.; Kothari, P.J.

    1984-01-01

    Many of the uniquely labelled synthetic precursors currently employed in the design of sophisticated radiolabelled compounds have their origins in the field of hot atom chemistry. Particularly, the development during the past few years of automated, on-line synthetic procedures which combine the nuclear reaction, hot atom and classical chemistry, and rapid purification methods has allowed the incorporation of useful radionuclides into suitable compounds of chemical and biochemical interest. The application of isotopically labelled methyl iodide, formaldehyde, and cyanide anion as synthetic intermediates in research involving human physiology and nuclear medicine, as well as their contributions to other scientific methodology, is reviewed. (author)

  5. Innovative method for carbon dioxide determination in human postmortem cardiac gas samples using headspace-gas chromatography–mass spectrometry and stable labeled isotope as internal standard

    Energy Technology Data Exchange (ETDEWEB)

    Varlet, V., E-mail: vincent.varlet@chuv.ch [Toxicology and Forensic Chemistry Unit, University Center of Legal Medicine Lausanne – Geneva, CH-1011 Lausanne (Switzerland); Smith, F. [Toxicology and Forensic Chemistry Unit, University Center of Legal Medicine Lausanne – Geneva, CH-1011 Lausanne (Switzerland); Froidmont, S. de; Dominguez, A.; Rinaldi, A. [Forensic Medicine Unit, University Center of Legal Medicine Lausanne – Geneva, CH-1011 Lausanne (Switzerland); Augsburger, M. [Toxicology and Forensic Chemistry Unit, University Center of Legal Medicine Lausanne – Geneva, CH-1011 Lausanne (Switzerland); Mangin, P.; Grabherr, S. [Forensic Medicine Unit, University Center of Legal Medicine Lausanne – Geneva, CH-1011 Lausanne (Switzerland)

    2013-06-19

    Graphical abstract: -- Highlights: •We developed a method for CO{sub 2} analysis in cardiac samples and quantification by {sup 13}CO{sub 2}. •This method was fully validated by accuracy profile. •We have applied this method to perform CO{sub 2} precise quantification for forensic applications. •Context of the death could be documented following CO{sub 2} concentrations. -- Abstract: A novel approach to measure carbon dioxide (CO{sub 2}) in gaseous samples, based on a precise and accurate quantification by {sup 13}CO{sub 2} internal standard generated in situ is presented. The main goal of this study was to provide an innovative headspace-gas chromatography–mass spectrometry (HS-GC–MS) method applicable in the routine determination of CO{sub 2}. The main drawback of the GC methods discussed in the literature for CO{sub 2} measurement is the lack of a specific internal standard necessary to perform quantification. CO{sub 2} measurement is still quantified by external calibration without taking into account analytical problems which can often occur considering gaseous samples. To avoid the manipulation of a stable isotope-labeled gas, we have chosen to generate in situ an internal labeled standard gas ({sup 13}CO{sub 2}) on the basis of the stoichiometric formation of CO{sub 2} by the reaction of hydrochloric acid (HCl) with sodium hydrogen carbonate (NaH{sup 13}CO{sub 3}). This method allows a precise measurement of CO{sub 2} concentration and was validated on various human postmortem gas samples in order to study its efficiency.

  6. Isotope labeling for NMR studies of macromolecular structure and interactions

    International Nuclear Information System (INIS)

    Wright, P.E.

    1994-01-01

    Implementation of biosynthetic methods for uniform or specific isotope labeling of proteins, coupled with the recent development of powerful heteronuclear multidimensional NMR methods, has led to a dramatic increase in the size and complexity of macromolecular systems that are now amenable to NMR structural analysis. In recent years, a new technology has emerged that combines uniform 13 C, 15 N labeling with heteronuclear multidimensional NMR methods to allow NMR structural studies of systems approaching 25 to 30 kDa in molecular weight. In addition, with the introduction of specific 13 C and 15 N labels into ligands, meaningful NMR studies of complexes of even higher molecular weight have become feasible. These advances usher in a new era in which the earlier, rather stringent molecular weight limitations have been greatly surpassed and NMR can begin to address many central biological problems that involve macromolecular structure, dynamics, and interactions

  7. Isotope labeling for NMR studies of macromolecular structure and interactions

    Energy Technology Data Exchange (ETDEWEB)

    Wright, P.E. [Scripps Research Institute, La Jolla, CA (United States)

    1994-12-01

    Implementation of biosynthetic methods for uniform or specific isotope labeling of proteins, coupled with the recent development of powerful heteronuclear multidimensional NMR methods, has led to a dramatic increase in the size and complexity of macromolecular systems that are now amenable to NMR structural analysis. In recent years, a new technology has emerged that combines uniform {sup 13}C, {sup 15}N labeling with heteronuclear multidimensional NMR methods to allow NMR structural studies of systems approaching 25 to 30 kDa in molecular weight. In addition, with the introduction of specific {sup 13}C and {sup 15}N labels into ligands, meaningful NMR studies of complexes of even higher molecular weight have become feasible. These advances usher in a new era in which the earlier, rather stringent molecular weight limitations have been greatly surpassed and NMR can begin to address many central biological problems that involve macromolecular structure, dynamics, and interactions.

  8. Characterization and relative quantification of phospholipids based on methylation and stable isotopic labeling[S

    Science.gov (United States)

    Cai, Tanxi; Shu, Qingbo; Liu, Peibin; Niu, Lili; Guo, Xiaojing; Ding, Xiang; Xue, Peng; Xie, Zhensheng; Wang, Jifeng; Zhu, Nali; Wu, Peng; Niu, Lili; Yang, Fuquan

    2016-01-01

    Phospholipids (PLs), one of the lipid categories, are not only the primary building blocks of cellular membranes, but also can be split to produce products that function as second messengers in signal transduction and play a pivotal role in numerous cellular processes, including cell growth, survival, and motility. Here, we present an integrated novel method that combines a fast and robust TMS-diazomethane-based phosphate derivatization and isotopic labeling strategy, which enables simultaneous profiling and relative quantification of PLs from biological samples. Our results showed that phosphate methylation allows fast and sensitive identification of the six major PL classes, including their lysophospholipid counterparts, under positive ionization mode. The isotopic labeling of endogenous PLs was achieved by deuterated diazomethane, which was generated through acid-catalyzed hydrogen/deuterium (H/D) exchange and methanolysis of TMS-diazomethane during the process of phosphate derivatization. The measured H/D ratios of unlabeled and labeled PLs, which were mixed in known proportions, indicated that the isotopic labeling strategy is capable of providing relative quantitation with adequate accuracy, reproducibility, and a coefficient of variation of 9.1%, on average. This novel method offers unique advantages over existing approaches and presents a powerful tool for research of PL metabolism and signaling. PMID:26733148

  9. Isotope separations using chromatographic methods

    International Nuclear Information System (INIS)

    Leseticky, L.

    1985-01-01

    A survey is given of chromatographic separations of compounds only differing in isotope composition. Isotope effects on physical properties which allow chromatographic separation (vapour tension, adsorption heat, partition coefficient) are very small, with the exception of the simplest molecules. Therefore, separation factors only assume the value of several per cent. From this ensues the necessity of using columns which are specially and very carefully prepared and have a separation efficiency of the order of 10 4 theoretical plates. Briefly discussed is liquid chromatography on ion exchangers which with a varied degree of success was used for separating simple inorganic compounds or ions. Ion exchange chromatography of amino acids labelled with tritium, and chromatography of tritium labelled steroids also provided only a certain degree of separation. A detailed analysis is presented of gas chromatography separation of various deuterium and tritium labelled low-molecular compounds, to which a number of studies has been devoted in the literature. Very promising is the method of complexation gas chromatography based on the reversible formation of a complex of the ligand (the compound being separated) and the compound of the (transition) metal as the steady-state phase. (author)

  10. Synthetic routes to some isotopically labelled intermediates for diterpenoid biosynthesis

    International Nuclear Information System (INIS)

    Dawson, R.M.; Godfrey, I.M.; Hogg, R.W.; Knox, J.R.

    1989-01-01

    The exo-15-hydrogen of ent-kaurene can be exchanged through a reversible ene reaction in a convenient and efficient procedure which has the potential for giving high specific activity 3 H-labelling. Copalol, the (Z)-double bond stereoisomer, and the allylic alcohol isomers ent-manool and ent-epimanool have been obtained through divergent synthetic pathways involving a 15,16-bisnor ketone intermediate. These pathways have also allowed the four compounds to be obtained with 14 C-labelling. A method, involving a Wittig reaction to form a vinyl bromide intermediate, has been developed for obtaining copalol, as the trityl ether derivative, with stereospecific isotopic labelling of one or the other of the hydrogens of the exocyclic methylene group. 27 refs., figs

  11. Tests of intestinal absorption using carbon-14-labeled isotopes

    International Nuclear Information System (INIS)

    Fromm, H.; Sarva, R.P.

    1983-01-01

    Beta radiation-emitting isotopes are being used increasingly in diagnostic gastroenterology for the study of absorption. The major reason for the popularity of radioisotopes is that their use is convenient for patient and physician alike. They often obviate naso- or orointestinal intubation and the collection, storage, and analysis of stool. The radioactivity used for the studies of digestive and absorptive processes is small and is not hazardous. In spite of the safety of the radiolabeled compounds, their use is restricted in children and pregnant women. Therefore, for most tests, promising alternative methods that make use of the stable isotope of carbon, /sup 13/C, instead of the radioactive /sup 14/C have been developed. The analysis of stable isotopes requires more sophisticated technology than that of radioactive compounds, however. Only a few centers presently are equipped and staffed to analyze stable isotopes on a routine basis. In contrast, the analysis of radioactive isotopes has become a routine procedure in almost ever major laboratory. The last decade has brought the development of several radioactive absorption tests. The clinically most useful tests relate to the study of bile acid, fat, lactose, and xylose absorption. All of these tests utilize the excretion rate of /sup 14/CO/sub 2/ in breath after ingestion of a /sup 14/C-labeled compound as a measure of the rate of its absorption or malabsorption

  12. 13C metabolic flux analysis: optimal design of isotopic labeling experiments.

    Science.gov (United States)

    Antoniewicz, Maciek R

    2013-12-01

    Measuring fluxes by 13C metabolic flux analysis (13C-MFA) has become a key activity in chemical and pharmaceutical biotechnology. Optimal design of isotopic labeling experiments is of central importance to 13C-MFA as it determines the precision with which fluxes can be estimated. Traditional methods for selecting isotopic tracers and labeling measurements did not fully utilize the power of 13C-MFA. Recently, new approaches were developed for optimal design of isotopic labeling experiments based on parallel labeling experiments and algorithms for rational selection of tracers. In addition, advanced isotopic labeling measurements were developed based on tandem mass spectrometry. Combined, these approaches can dramatically improve the quality of 13C-MFA results with important applications in metabolic engineering and biotechnology. Copyright © 2013 Elsevier Ltd. All rights reserved.

  13. Investigation into reaction of heterogenous isotopic exchange with gaseoUs tritium in solution for preparation labelled lipid compounds

    International Nuclear Information System (INIS)

    Shevchenko, V.P.; Myasoedov, N.F.

    1983-01-01

    The applicability of the method of heterogeneous catalytic isotopic exchange with gaseous tritium in the solution for the production of labelled lipide preparations is studied. Labelled saturated and unsaturated aliphatic acids, prostaglandins, phospholipides and sphingolipides are prepared

  14. Isotope-labelled folic acid derivatives

    International Nuclear Information System (INIS)

    Lewin, N.; Wong, E.T.

    1976-01-01

    The suggestion deals with the production of folic acid derivatives suitable as indicators or tracers for analyses of serum folates. These folic acid derivatives contain folic acid which is bound by one or both carboxyl groups to the amino nitrogen of compounds such as, e.g., tyramine, glycyl tyrosine, tyrosine, or the methyl ester of tyrosine. The derivative obtained can be substituted by a gamma emitter, e.g. the iodine isotope I 125. The radioactive derivative is used in the method for the competitive protein bonding to determine endogenic folates in the serum. (UWI) [de

  15. Stable isotope labeling strategy based on coding theory

    Energy Technology Data Exchange (ETDEWEB)

    Kasai, Takuma; Koshiba, Seizo; Yokoyama, Jun; Kigawa, Takanori, E-mail: kigawa@riken.jp [RIKEN Quantitative Biology Center (QBiC), Laboratory for Biomolecular Structure and Dynamics (Japan)

    2015-10-15

    We describe a strategy for stable isotope-aided protein nuclear magnetic resonance (NMR) analysis, called stable isotope encoding. The basic idea of this strategy is that amino-acid selective labeling can be considered as “encoding and decoding” processes, in which the information of amino acid type is encoded by the stable isotope labeling ratio of the corresponding residue and it is decoded by analyzing NMR spectra. According to the idea, the strategy can diminish the required number of labelled samples by increasing information content per sample, enabling discrimination of 19 kinds of non-proline amino acids with only three labeled samples. The idea also enables this strategy to combine with information technologies, such as error detection by check digit, to improve the robustness of analyses with low quality data. Stable isotope encoding will facilitate NMR analyses of proteins under non-ideal conditions, such as those in large complex systems, with low-solubility, and in living cells.

  16. Stable isotope labeling strategy based on coding theory

    International Nuclear Information System (INIS)

    Kasai, Takuma; Koshiba, Seizo; Yokoyama, Jun; Kigawa, Takanori

    2015-01-01

    We describe a strategy for stable isotope-aided protein nuclear magnetic resonance (NMR) analysis, called stable isotope encoding. The basic idea of this strategy is that amino-acid selective labeling can be considered as “encoding and decoding” processes, in which the information of amino acid type is encoded by the stable isotope labeling ratio of the corresponding residue and it is decoded by analyzing NMR spectra. According to the idea, the strategy can diminish the required number of labelled samples by increasing information content per sample, enabling discrimination of 19 kinds of non-proline amino acids with only three labeled samples. The idea also enables this strategy to combine with information technologies, such as error detection by check digit, to improve the robustness of analyses with low quality data. Stable isotope encoding will facilitate NMR analyses of proteins under non-ideal conditions, such as those in large complex systems, with low-solubility, and in living cells

  17. Synthesis of 14C-labeled and stable isotope-labeled CGS 16617

    International Nuclear Information System (INIS)

    Chaudhuri, N.K.; Markus, B.; Sung Mingsang

    1988-01-01

    The synthesis of a 14 C-labeled and two stable isotope-labeled analogs of CGS 16617 is described. The synthetic method involved the preparation of tetrahydro-3-bromo-1-benzazepin-2-one, labeled with a 14 C or four deuterium atoms, followed by introduction of two side chains at 1- and 3-positions. The labeled bromobenzazepinones were prepared by Beckmann rearrangement of bromo-oximes of α-tetralones, obtained by cyclization of labeled benzenebutanoic acids. The 14 C-labeled acid was prepared by hydrolysis of the nitrile, prepared by reaction of 3-bromopropylbenzene and K 14 CN. The tetradeutero acid was prepared from ethyl phenylpropynoate by catalytic reduction of the triple bond with deuterium gas, followed by reduction of the deuterated ester with lithium aluminium hydride and conversion of the resulting alcohol into the carboxylic acid. The acetic acid side chain was introduced by N-alkylation with ethyl bromoacetate or ethyl bromoacetate-1, 2- 13 C 2 followed by hydrolysis, and the L-lysine side chain, by reaction with L-(-)-3-amino-ε-caprolactam followed by hydrolysis of the caprolactam ring. (author)

  18. Method for separating isotopes

    International Nuclear Information System (INIS)

    Jepson, B.E.

    1975-01-01

    Isotopes are separated by contacting a feed solution containing the isotopes with a cyclic polyether wherein a complex of one isotope is formed with the cyclic polyether, the cyclic polyether complex is extracted from the feed solution, and the isotope is thereafter separated from the cyclic polyether

  19. Stereoselective synthesis of stable-isotope-labeled amino acids

    Energy Technology Data Exchange (ETDEWEB)

    Unkefer, C.J.; Martinez, R.A.; Silks, L.A. III [Los Alamos National Laboratory, NM (United States); Lodwig, S.N. [Centralia College, WA (United States)

    1994-12-01

    For magnetic resonance and vibrational spectroscopies to reach their full potential, they must be used in combination with sophisticated site-specific stable isotope labeling of biological macromolecules. Labeled amino acids are required for the study of the structure and function of enzymes and proteins. Because there are 20 common amino acids, each with its own distinguishing chemistry, they remain a synthetic challenge. The Oppolzer chiral auxiliary provides a general tool with which to approach the synthesis of labeled amino acids. By using the Oppolzer auxiliary, amino acids can be constructed from several small molecules, which is ideal for stable isotope labeling. In addition to directing the stereochemistry at the {alpha}-carbon, the camphorsultam can be used for stereo-specific isotope labeling at prochiral centers in amino acids. By using the camphorsultam auxiliary we have the potential to synthesize virtually any isotopomer of all of the common amino acids.

  20. Stereoselective synthesis of stable-isotope-labeled amino acids

    International Nuclear Information System (INIS)

    Unkefer, C.J.; Martinez, R.A.; Silks, L.A. III; Lodwig, S.N.

    1994-01-01

    For magnetic resonance and vibrational spectroscopies to reach their full potential, they must be used in combination with sophisticated site-specific stable isotope labeling of biological macromolecules. Labeled amino acids are required for the study of the structure and function of enzymes and proteins. Because there are 20 common amino acids, each with its own distinguishing chemistry, they remain a synthetic challenge. The Oppolzer chiral auxiliary provides a general tool with which to approach the synthesis of labeled amino acids. By using the Oppolzer auxiliary, amino acids can be constructed from several small molecules, which is ideal for stable isotope labeling. In addition to directing the stereochemistry at the α-carbon, the camphorsultam can be used for stereo-specific isotope labeling at prochiral centers in amino acids. By using the camphorsultam auxiliary we have the potential to synthesize virtually any isotopomer of all of the common amino acids

  1. Method for separating isotopes

    International Nuclear Information System (INIS)

    Schlenker, R.F.

    1978-01-01

    A vortex tube for separating isotopes is described. A gas mixture containing the isotopic molecules enters the vortex tube under pressure and is separated into a hot discharge flow stream and a cold discharge flow stream. The hot discharge is enriched in lighter isotopic molecules whereas the cold discharge flow stream is enriched in the heavier isotopic molecules. The vortex tube can be used in a single stage or multistage isotope separation apparatus

  2. Production of isotopically labeled heterologous proteins in non-E. coli prokaryotic and eukaryotic cells

    International Nuclear Information System (INIS)

    Takahashi, Hideo; Shimada, Ichio

    2010-01-01

    The preparation of stable isotope-labeled proteins is necessary for the application of a wide variety of NMR methods, to study the structures and dynamics of proteins and protein complexes. The E. coli expression system is generally used for the production of isotope-labeled proteins, because of the advantages of ease of handling, rapid growth, high-level protein production, and low cost for isotope-labeling. However, many eukaryotic proteins are not functionally expressed in E. coli, due to problems related to disulfide bond formation, post-translational modifications, and folding. In such cases, other expression systems are required for producing proteins for biomolecular NMR analyses. In this paper, we review the recent advances in expression systems for isotopically labeled heterologous proteins, utilizing non-E. coli prokaryotic and eukaryotic cells.

  3. Isotope separation method and apparatus

    International Nuclear Information System (INIS)

    Lyon, R.K.; Eisner, P.N.; Thomas, W.R.L.

    1980-01-01

    A method and apparatus are specified for separating a mixture of isotopes present in a compound, preferably a gaseous compound, into two or more parts in each of which the abundances of the isotopes differ from the natural abundances of the isotopes in the compound. The invention particularly relates to carrying out a laser induced, isotopically selective conversion of gaseous molecules in such a manner as to achieve more than one stage of isotope separation along the length of the laser beam. As an example, the invention is applied to the separation of the isotopes of uranium in UF 6 , in which either the U-235 or U-238 isotope is selectively excited by means of irradiation from an infrared laser, and the selectively excited isotope converted into a product that can be recovered from UF 6 by one of a variety of methods that are described. (U.K.)

  4. Method of isotope separation

    Energy Technology Data Exchange (ETDEWEB)

    Lyon, R K

    1975-05-22

    Isotopes of a gaseous compound can be separated by multi-infrared photoabsorption which follows a selective dissociation of the excited molecules by single photon absorption of photons of visible or UV radiation. The process involves three steps. Firstly, the molecules to be separated are irradiated with a high-energy IR laser, whereby the molecules of the compound containing the lighter isotopes are preferably excited. They are then irradiated by a second laser with UV or visible light whose frequency of radiation brings the excited molecules into a form in which they can be separated from the non-excited molecules. The third step is the reformation of the substances according to known methods. A power density of at least 10/sup 4/ watt/cm/sup 2/ per torr gas pressure with an irradiation time of 10/sup -10/ to 5 x 10/sup -5/ seconds in the presence of a second gas with at least 5 times higher partial pressure is necessary for the IR radiation. The method may be used for UF/sub 6/ for which an example is given here.

  5. Custom synthesis of isotope-labelled Apis mellifera Pheromone

    International Nuclear Information System (INIS)

    Conanan, Aida P.; Cortes, Nicole Marie A.; Daguno, Cristel Lyn R.; Templonuevo, Jose Angelo A.; Sucgang, Raymond J.

    2012-01-01

    The object of this study is to determine the optimum conditions for the synthesis of the isotope-labelled isopentyl acetate. Isopentyl acetate is widely used as a raw material in industries, in syntheses, and is utilized as a sex attractant (pheromone) by the bee species, Apis mellifera. The isotope labelling of isopentyl acetate will allow tracking of the fate and movement of the isopentyl acetate in the environment, in chemical transformations, and in biological systems. Esterification by alcoholysis of acetic acid was optimized for the preparation of Carbon-14( 14 C)-labelled isopentyl acetate from 14 C-labelled acetic acid and isoamyl alcohol. The different conditions studied were: (1) The effects of acid catalysis and/or reflux on the incorporation and retention of the isotope label on the product. The efficiency of label incorporation and retention was determined through the beta radioactivity of Carbon 14 in each of the synthetic constructs. Determination of the beta radioactivity concentration of 14 C in the isopentyl acetate product was done using low level liquid scintillation spectrometry. Each of the synthetic products was mixed with Ultima Gold scintillation cocktail in a low potassium glass scintillation vial, and analysed in a low-level Wallac 1414 scintillation counter. The application of catalysis without reflux resulted in the highest yield (35%). The same condition also resulted in the highest abundance of carbon isotope label with 2.40 Bequerels per cubic centimetre, Bq/cc (measurement unit for radioactivity). (author)

  6. Linerless label device and method

    KAUST Repository

    Binladen, Abdulkari

    2016-01-14

    This apparatus and method for applying a linerless label to an end user product includes a device with a printer for printing on a face surface of a linerless label, and a release coat applicator for applying a release coat to the face surface of the label; another device including an unwinder unit (103) to unwind a roll of printed linerless label; a belt (108); a glue applicator (102) for applying glue to the belt; a nip roller (106) for contacting and applying pressure to the face surface of the linerless label such that the glue on the belt transfers to the back surface of the linerless label; at least one slitting knife 105) positioned downstream the belt and a rewinder unit (104) positioned downstream the slitting knife; and a third device which die cuts and applies the linerless label to an end user object.

  7. A new combined method of stable isotope-labeling derivatization-ultrasound-assisted dispersive liquid-liquid microextraction for the determination of neurotransmitters in rat brain microdialysates by ultra high performance liquid chromatography tandem mass spectrometry.

    Science.gov (United States)

    Zheng, Longfang; Zhao, Xian-En; Zhu, Shuyun; Tao, Yanduo; Ji, Wenhua; Geng, Yanling; Wang, Xiao; Chen, Guang; You, Jinmao

    2017-06-01

    In this work, for the first time, a new hyphenated technique of stable isotope-labeling derivatization-ultrasound-assisted dispersive liquid-liquid microextraction has been developed for the simultaneous determination of monoamine neurotransmitters (MANTs) and their biosynthesis precursors and metabolites. The developed method was based on ultra high performance liquid chromatography tandem mass spectrometry detection using multiple-reaction monitoring mode. A pair of mass spectrometry sensitizing reagents, d 0 -10-methyl-acridone-2-sulfonyl chloride and d 3 -10-methyl-acridone-2-sulfonyl chloride, as stable isotope probes was utilized to facilely label neurotransmitters, respectively. The heavy labeled MANTs standards were prepared and used as internal standards for quantification to minimize the matrix effects in mass spectrometry analysis. Low toxic bromobenzene (extractant) and acetonitrile (dispersant) were utilized in microextraction procedure. Under the optimized conditions, good linearity was observed with the limits of detection (S/N>3) and limits of quantification (S/N>10) in the range of 0.002-0.010 and 0.015-0.040nmol/L, respectively. Meanwhile, it also brought acceptable precision (4.2-8.8%, peak area RSDs %) and accuracy (recovery, 96.9-104.1%) results. This method was successfully applied to the simultaneous determination of monoamine neurotransmitters and their biosynthesis precursors and metabolites in rat brain microdialysates of Parkinson's disease and normal rats. This provided a new method for the neurotransmitters related studies in the future. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Linerless label device and method

    KAUST Repository

    Binladen, Abdulkari

    2016-01-01

    This apparatus and method for applying a linerless label to an end user product includes a device with a printer for printing on a face surface of a linerless label, and a release coat applicator for applying a release coat to the face surface

  9. Basic methods of isotope analysis

    International Nuclear Information System (INIS)

    Ochkin, A.V.; Rozenkevich, M.B.

    2000-01-01

    The bases of the most applied methods of the isotope analysis are briefly presented. The possibilities and analytical characteristics of the mass-spectrometric, spectral, radiochemical and special methods of the isotope analysis, including application of the magnetic resonance, chromatography and refractometry, are considered [ru

  10. Reconstitution of Nucleosomes with Differentially Isotope-labeled Sister Histones.

    Science.gov (United States)

    Liokatis, Stamatios

    2017-03-26

    Asymmetrically modified nucleosomes contain the two copies of a histone (sister histones) decorated with distinct sets of Post-translational Modifications (PTMs). They are newly identified species with unknown means of establishment and functional implications. Current analytical methods are inadequate to detect the copy-specific occurrence of PTMs on the nucleosomal sister histones. This protocol presents a biochemical method for the in vitro reconstitution of nucleosomes containing differentially isotope-labeled sister histones. The generated complex can be also asymmetrically modified, after including a premodified histone pool during refolding of histone subcomplexes. These asymmetric nucleosome preparations can be readily reacted with histone-modifying enzymes to study modification cross-talk mechanisms imposed by the asymmetrically pre-incorporated PTM using nuclear magnetic resonance (NMR) spectroscopy. Particularly, the modification reactions in real-time can be mapped independently on the two sister histones by performing different types of NMR correlation experiments, tailored for the respective isotope type. This methodology provides the means to study crosstalk mechanisms that contribute to the formation and propagation of asymmetric PTM patterns on nucleosomal complexes.

  11. Method for separating krypton isotopes

    International Nuclear Information System (INIS)

    Porter, J.T.

    1980-01-01

    Methods and apparatus for separating krypton isotopes utilizing low temperature selective infrared excitation of 85krypton difluoride in an isotopic compound mixture. Multiphoton ir excitation and uv excitation techniques are used, as well as cryogenic matrix isolation and inert buffer gas isolation techniques

  12. Heating Isotopically Labeled Bernal Stacked Graphene: A Raman Spectroscopy Study

    Czech Academy of Sciences Publication Activity Database

    Ek Weis, Johan; da Costa, Sara; Frank, Otakar; Kalbáč, Martin

    2014-01-01

    Roč. 5, č. 3 (2014), s. 549-554 ISSN 1948-7185 R&D Projects: GA MŠk LL1301 Institutional support: RVO:61388955 Keywords : Bernal * graphene * isotopic labeling Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 7.458, year: 2014

  13. Performance of isobaric and isotopic labeling in quantitative plant proteomics

    DEFF Research Database (Denmark)

    Nogueira, Fábio C S; Palmisano, Giuseppe; Schwämmle, Veit

    2012-01-01

    , and quantitation. In the present work, we have used LC-MS to compare an isotopic (ICPL) and isobaric (iTRAQ) chemical labeling technique to quantify proteins in the endosperm of Ricinus communis seeds at three developmental stages (IV, VI, and X). Endosperm proteins of each stage were trypsin-digested in...

  14. Method to separate isotopes

    International Nuclear Information System (INIS)

    Coenen, H.; Neuschuetz, D.

    1980-01-01

    An extraction by means of supercritical pure gases (e.g. CD 2 ) or gas mixtures is proposed to separate isotopes, especially H/D, Li-6/Li-7, and U-235/U-238, and water or benzol is used as entrainer. The extraction shall be carried out at pressure rates of about 350 bar above the critical pressure with the temperature being by up to 100 0 C above the critical temperature. A NaCl-solution and the change of the isotopic ratio Cl-35/Cl-37 are investigated for example purpose. (UWI) [de

  15. Custom isotope-labelling of apis mellifera pheromone

    International Nuclear Information System (INIS)

    Conanan, Aida P.; Cortes, Nicole Marie A.; Daguno, Cristel Lyn R.; Templonuevo, Jose Angelo A.; Sucgang, Raymond J.

    2012-01-01

    The object of this study is to determine the optimum conditions for the synthesis of isotope-labelled isoamyl acetate. Esterification by alcoholysis of acetic acid was optimized for the preparation of Carbon - 14 ( 14 C)-labelled isopentyl acetate from 14 C-labelled acetic acid and isopentyl alcohol. The optimization procedure defined the effects of catalysis, reflux time, and temperature. The application of catalysis without reflux resulted to the highest yield (40%); the same condition also resulted to the highest abundance of carbon isotope label with 2.40 disintegrations per minute per cubic centimetre, DPM/cc (measurement unit for radioactivity). Determination of the beta radioactivity concentration of 14 C in the isopentyl acetate product was done using low level liquid scintillation spectrometry. Each of the synthetic constructs was mixed with Ultima Gold scintillation cocktail in a glass scintillation vial, and analysed in a low-level Wallac 1414 scintillation counter. Samples were counted for 2 hours in a chamber temperature maintained at 14 degree centegrade. The catalysed reaction without reflux was established to be the most efficient scheme for the radiolabelling. The radiolabelled isoamyl acetate can give way to the synthesis of more complex substances which can be then tracked when they are introduced to a system through the carbon isotope label. (author)

  16. Determination of photooxygenation products of rotenone with isotope dilution method

    International Nuclear Information System (INIS)

    Chubachi, Mitsuo; Hamada, Masayuki

    1975-01-01

    When rotenone dissolved in certain solvent was photochemically oxidized, rotenolones, dehydrorotenone and rotenonone were obtained as main products. In order to determine the quantitative yields of these compounds in photooxygenation products, four compounds mentioned above were labeled with carbon-14 and the isotope dilution method by these labeled compounds was applied to the product analysis. (auth.)

  17. Availability of phosphorus in cow slurry using isotopic labelling technique

    International Nuclear Information System (INIS)

    Pongsakul, P.; Bertelsen, F.; Gissel-Nielsen, G.

    1988-01-01

    A pot experiment was conducted to evaluate the influence of cow slurry on P uptake by corn and to estimate the readily available P in the slurry by using an isotopic labelling techique. Water-soluble P in soil was increased and isotopic equilibrium of available P was attained after labelled slurry was mixed thoroughly throughout the soil. Labelled slurry applied at planting increased the P uptake by corn, whereas the same amount applied one week before harvest did not affect the P uptake. It was estimated that 46-54% of the total P uptake in plants is derived from the slurry. The readily available P (the L-value) in the slurry was at least 26 mg/kg which equals 3.7% of the total P. (author)

  18. Escherichia coli cell-free protein synthesis and isotope labeling of mammalian proteins.

    Science.gov (United States)

    Terada, Takaho; Yokoyama, Shigeyuki

    2015-01-01

    This chapter describes the cell-free protein synthesis method, using an Escherichia coli cell extract. This is a cost-effective method for milligram-scale protein production and is particularly useful for the production of mammalian proteins, protein complexes, and membrane proteins that are difficult to synthesize by recombinant expression methods, using E. coli and eukaryotic cells. By adjusting the conditions of the cell-free method, zinc-binding proteins, disulfide-bonded proteins, ligand-bound proteins, etc., may also be produced. Stable isotope labeling of proteins can be accomplished by the cell-free method, simply by using stable isotope-labeled amino acid(s) in the cell-free reaction. Moreover, the cell-free protein synthesis method facilitates the avoidance of stable isotope scrambling and dilution over the recombinant expression methods and is therefore advantageous for amino acid-selective stable isotope labeling. Site-specific stable isotope labeling is also possible with a tRNA molecule specific to the UAG codon. By the cell-free protein synthesis method, coupled transcription-translation is performed from a plasmid vector or a PCR-amplified DNA fragment encoding the protein. A milligram quantity of protein can be produced with a milliliter-scale reaction solution in the dialysis mode. More than a thousand solution structures have been determined by NMR spectroscopy for uniformly labeled samples of human and mouse functional domain proteins, produced by the cell-free method. Here, we describe the practical aspects of mammalian protein production by the cell-free method for NMR spectroscopy. © 2015 Elsevier Inc. All rights reserved.

  19. Evaluation of a Method for Nitrotyrosine Site Identification and Relative Quantitation Using a Stable Isotope-Labeled Nitrated Spike-In Standard and High Resolution Fourier Transform MS and MS/MS Analysis

    Directory of Open Access Journals (Sweden)

    Kent W. Seeley

    2014-04-01

    Full Text Available The overproduction of reactive oxygen and nitrogen species (ROS and RNS can have deleterious effects in the cell, including structural and possible activity-altering modifications to proteins. Peroxynitrite is one such RNS that can result in a specific protein modification, nitration of tyrosine residues to form nitrotyrosine, and to date, the identification of nitrotyrosine sites in proteins continues to be a major analytical challenge. We have developed a method by which 15N-labeled nitrotyrosine groups are generated on peptide or protein standards using stable isotope-labeled peroxynitrite (O15NOO−, and the resulting standard is mixed with representative samples in which nitrotyrosine formation is to be measured by mass spectrometry (MS. Nitropeptide MS/MS spectra are filtered using high mass accuracy Fourier transform MS (FTMS detection of the nitrotyrosine immonium ion. Given that the nitropeptide pair is co-isolated for MS/MS fragmentation, the nitrotyrosine immonium ions (at m/z = 181 or 182 can be used for relative quantitation with negligible isotopic interference at a mass resolution of greater than 50,000 (FWHM, full width at half-maximum. Furthermore, the standard potentially allows for the increased signal of nitrotyrosine-containing peptides, thus facilitating selection for MS/MS in a data-dependent mode of acquisition. We have evaluated the methodology in terms of nitrotyrosine site identification and relative quantitation using nitrated peptide and protein standards.

  20. Mass spectrometric studies of stable isotope-labelled carboxylic acid derivatives

    International Nuclear Information System (INIS)

    Andersson, B.Aa.; Dinger, F.; Dinh-Nguyen, N.

    1975-01-01

    Low resolution mass spectra of deuterium and carbon-13 labelled fatty acid pyrrolidides are discussed. The simple fragmentation pattern of pyrrolidides makes them superior to other derivatives, regarding location of isotopes. Deuteriation of ethylenic fatty acid pyrrolidides therefore seems to be an improved method to locate carbon-carbon double bonds by mass spectrometry. (author)

  1. A method for the determination of bacterial spore DNA content based on isotopic labelling, spore germination and diphenylamine assay; ploidy of spores of several Bacillus species

    International Nuclear Information System (INIS)

    Hauser, P.M.; Karamata, D.

    1992-01-01

    A reliable method for measuring the spore DNA content, based on radioactive DNA labelling, spore germination in absence of DNA replication and diphenylamine assay, was developed. The accuracy of the method, within 10 - 15%, is adequate for determining the number of chromosomes per spore, provided that the genome size is known. B subtilis spores were shown to be invariably monogenomic, while those of larger bacilli Bacillus megaterium, Bacillus cereus and Bacillus thuringiensis, often, if not invariably, contain two genomes. Attempts to modify the spore DNA content of B subtilis by altering the richness of the sporulation medium, the sporulation conditions (liquid or solid medium), or by mutation, were apparently unsuccessful. An increase of spore size with medium richness, not accompanied by an increase in DNA content, was observed. The implication of the apparently species-specific spore ploidy and the influence of the sporulation conditions on spore size and shape are discussed

  2. Radioisotope methods for leucocyte labelling

    International Nuclear Information System (INIS)

    Kostadinova, I.; Kovacheva, S.

    1988-01-01

    A review is made of the labelling methods with the following tracers: 3 H-thymidine, 32 P-DP, 111 In (oxine, tropolon, acetylacetone, MERC), 99m Tc (reduced 99m Tc, lypophyl 99m Tc-complexes and 99m Tc-colloids). The main diagnosis areas are mentioned: abdominal abscesses and inflammations, inflammation foci of skeleton or of implanted prosthesis; acute myocardial infarction, bacterial endocarditis, rejection of kydney transplantations or vascular grafts. It is concluded that labelled leucocytes are very reliable for noninvasive diagnosis of inflammation foci with unclear localization

  3. Radioisotope methods for leucocyte labelling

    Energy Technology Data Exchange (ETDEWEB)

    Kostadinova, I; Kovacheva, S [Meditsinska Akademiya, Sofia (Bulgaria). Katedra po Rentgenologiya i Radiologiya

    1988-01-01

    A review is made of the labelling methods with the following tracers: {sup 3}H-thymidine, {sup 32}P-DP, {sup 111}In (oxine, tropolon, acetylacetone, MERC), {sup 99m}Tc (reduced {sup 99m}Tc, lypophyl {sup 99m}Tc-complexes and {sup 99m}Tc-colloids). The main diagnosis areas are mentioned: abdominal abscesses and inflammations, inflammation foci of skeleton or of implanted prosthesis; acute myocardial infarction, bacterial endocarditis, rejection of kydney transplantations or vascular grafts. It is concluded that labelled leucocytes are very reliable for noninvasive diagnosis of inflammation foci with unclear localization.

  4. Isotope separation apparatus and method

    International Nuclear Information System (INIS)

    Feldman, B. J.

    1985-01-01

    The invention relates to an improved method and apparatus for laser isotope separation by photodeflection. A molecular beam comprising at least two isotopes to be separated intersects, preferably substantially perpendicular to one broad side of the molecular beam, with a laser beam traveling in a first direction. The laser beam is reflected back through the molecular beam, preferably in a second direction essentially opposite to the first direction. Because the molecules in the beam occupy various degenerate energy levels, if the laser beam comprises chirped pulses comprising selected wavelengths, the laser beam will very efficiently excite substantially all unexcited molecules and will cause stimulated emission of substantially all excited molecules of a selected one of the isotopes in the beam which such pulses encounter. Excitation caused by first direction chirped pulses moves molecules of the isotope excited thereby in the first direction. Stimulated emission of excited molecules of the isotope is brought about by returning chirped pulses traveling in the second direction. Stimulated emission moves emitting molecules in a direction opposite to the photon emitted. Because emitted photons travel in the second direction, emitting molecules move in the first direction. Substantial molecular movement of essentially all the molecules containing the one isotope is accomplished by a large number of chirped pulse-molecule interactions. A beam corer collects the molecules in the resulting enriched divergent portions of the beam

  5. Sensing site-specific structural characteristics and chirality using vibrational circular dichroism of isotope labeled peptides.

    Science.gov (United States)

    Keiderling, Timothy A

    2017-12-01

    Isotope labeling has a long history in chemistry as a tool for probing structure, offering enhanced sensitivity, or enabling site selection with a wide range of spectroscopic tools. Chirality sensitive methods such as electronic circular dichroism are global structural tools and have intrinsically low resolution. Consequently, they are generally insensitive to modifications to enhance site selectivity. The use of isotope labeling to modify vibrational spectra with unique resolvable frequency shifts can provide useful site-specific sensitivity, and these methods have been recently more widely expanded in biopolymer studies. While the spectral shifts resulting from changes in isotopic mass can provide resolution of modes from specific parts of the molecule and can allow detection of local change in structure with perturbation, these shifts alone do not directly indicate structure or chirality. With vibrational circular dichroism (VCD), the shifted bands and their resultant sign patterns can be used to indicate local conformations in labeled biopolymers, particularly if multiple labels are used and if their coupling is theoretically modeled. This mini-review discusses selected examples of the use of labeling specific amides in peptides to develop local structural insight with VCD spectra. © 2017 Wiley Periodicals, Inc.

  6. Modern spectrometric methods for the analysis of labelled compounds

    International Nuclear Information System (INIS)

    Kaspersen, F.M.; Funke, C.W.; Wagenaars, G.N.; Jacobs, P.L.

    1988-01-01

    A proper analysis of chemical compounds should give information about the chemical identity (not only the structure but also enantiomeric form), the chemical purity and chemical composition (e.g. giving information about counter-ions, solvents of crystallization). For labelled compounds information is also needed about isotopic purity (defined as the % of isotope present in the compound), the position/distribution of the isotope in the molecule and degree of labelling/specific activity. In the past ten years the possibilities for spectrometric analyses of labelled compounds have increased enormously and this chapter will give an overview of these methods with the exception of (radio)chromatography that will be dealt with in another chapter. (author)

  7. Isotope separation apparatus and method

    International Nuclear Information System (INIS)

    Cotter, T.P.

    1982-01-01

    The invention relates to a method and apparatus for laser isotope separation by photodeflection. A molecular beam comprising at least two isotopes to be separated intersects, preferable substantially perpendicular to one broad side of the molecular beam, with a laser beam traveling in a first direction. The laser beam is reflected back through the molecular beam, preferably in a second direction essentially opposite to the first direction. The laser beam comprises pi-pulses of a selected wavelength which excite unexcited molecules, or cause stimulated emission of excited molecules of one of the isotopes. Excitation caused by first direction pi-pulses moves molecules of the isotope excited thereby in the first direction. Stimulated emission of excited molecules of the isotope is brought about by returning pi-pulses traveling in the second direction. Stimulated emission moves emitting molecules in a direction opposite to the photon emitted. Because emitted photons travel in the second direction, emitting molecules move in the first direction. Substantial molecular movement is accomplished by a large number of pi-pulse-molecule interactions. A beam corer collects the molecules in the resulting enriched divergent portions of the beam

  8. Stable isotopic labeling-based quantitative targeted glycomics (i-QTaG).

    Science.gov (United States)

    Kim, Kyoung-Jin; Kim, Yoon-Woo; Kim, Yun-Gon; Park, Hae-Min; Jin, Jang Mi; Hwan Kim, Young; Yang, Yung-Hun; Kyu Lee, Jun; Chung, Junho; Lee, Sun-Gu; Saghatelian, Alan

    2015-01-01

    Mass spectrometry (MS) analysis combined with stable isotopic labeling is a promising method for the relative quantification of aberrant glycosylation in diseases and disorders. We developed a stable isotopic labeling-based quantitative targeted glycomics (i-QTaG) technique for the comparative and quantitative analysis of total N-glycans using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). We established the analytical procedure with the chemical derivatizations (i.e., sialic acid neutralization and stable isotopic labeling) of N-glycans using a model glycoprotein (bovine fetuin). Moreover, the i-QTaG using MALDI-TOF MS was evaluated with various molar ratios (1:1, 1:2, 1:5) of (13) C6 /(12) C6 -2-aminobenzoic acid-labeled glycans from normal human serum. Finally, this method was applied to direct comparison of the total N-glycan profiles between normal human sera (n = 8) and prostate cancer patient sera (n = 17). The intensities of the N-glycan peaks from i-QTaG method showed a good linearity (R(2) > 0.99) with the amount of the bovine fetuin glycoproteins. The ratios of relative intensity between the isotopically 2-AA labeled N-glycans were close to the theoretical molar ratios (1:1, 1:2, 1:5). We also demonstrated that the up-regulation of the Lewis antigen (~82%) in sera from prostate cancer patients. In this proof-of-concept study, we demonstrated that the i-QTaG method, which enables to achieve a reliable comparative quantitation of total N-glycans via MALDI-TOF MS analysis, has the potential to diagnose and monitor alterations in glycosylation associated with disease states or biotherapeutics. © 2015 American Institute of Chemical Engineers.

  9. Studies of phosphorus-containing fertilizer uptake in soils by 32P isotope labelling

    International Nuclear Information System (INIS)

    Fueleky, Gyoergy; Osztoics, Andrasne; Papne Kranitz, Erzsebet

    1983-01-01

    Breeding experiments were carried out with rye-grass (Lolium perenne L.) on two soil types to determine the plant uptake of phosphorus from naturally occuring element and from that added to the soil by superphosphate fertilizers. 32 P isotope labelling and radiometric measuring method were applied. In addition to the determination of phosphorus uptake, the phosphorus contents of the soil from its natural stock and from the fertilizer for both soil types can be determined by this method. (A.L.)

  10. Raman spectroscopy of isotopically labeled two-layer graphene

    Czech Academy of Sciences Publication Activity Database

    Kalbáč, Martin; Kong, J.; Kavan, Ladislav; Dresselhaus, M. S.

    2012-01-01

    Roč. 249, č. 12 (2012), s. 2500-2502 ISSN 0370-1972 R&D Projects: GA AV ČR IAA400400911; GA AV ČR IAA400400804; GA AV ČR KAN200100801; GA MŠk ME09060; GA ČR GAP204/10/1677; GA ČR GBP208/12/G016; GA ČR(CZ) GAP208/12/1062 Institutional support: RVO:61388955 Keywords : electrochemical doping * isotope labeling * graphene Subject RIV: CG - Electrochemistry Impact factor: 1.489, year: 2012

  11. Copper-catalyzed oxidative desulfurization-oxygenation of thiocarbonyl compounds using molecular oxygen: an efficient method for the preparation of oxygen isotopically labeled carbonyl compounds.

    Science.gov (United States)

    Shibahara, Fumitoshi; Suenami, Aiko; Yoshida, Atsunori; Murai, Toshiaki

    2007-06-21

    A novel copper-catalyzed oxidative desulfurization reaction of thiocarbonyl compounds, using molecular oxygen as an oxidant and leading to formation of carbonyl compounds, has been developed, and the utility of the process is demonstrated by its application to the preparation of a carbonyl-18O labeled sialic acid derivative.

  12. Quantitative O-glycomics based on improvement of the one-pot method for nonreductive O-glycan release and simultaneous stable isotope labeling with 1-(d0/d5)phenyl-3-methyl-5-pyrazolone followed by mass spectrometric analysis.

    Science.gov (United States)

    Wang, Chengjian; Zhang, Ping; Jin, Wanjun; Li, Lingmei; Qiang, Shan; Zhang, Ying; Huang, Linjuan; Wang, Zhongfu

    2017-01-06

    Rapid, simple and versatile methods for quantitative analysis of glycoprotein O-glycans are urgently required for current studies on protein O-glycosylation patterns and the search for disease O-glycan biomarkers. Relative quantitation of O-glycans using stable isotope labeling followed by mass spectrometric analysis represents an ideal and promising technique. However, it is hindered by the shortage of reliable nonreductive O-glycan release methods as well as the too large or too small inconstant mass difference between the light and heavy isotope form derivatives of O-glycans, which results in difficulties during the recognition and quantitative analysis of O-glycans by mass spectrometry. Herein we report a facile and versatile O-glycan relative quantification strategy, based on an improved one-pot method that can quantitatively achieve nonreductive release and in situ chromophoric labeling of intact mucin-type O-glycans in one step. In this study, the one-pot method is optimized and applied for quantitative O-glycan release and tagging with either non-deuterated (d 0 -) or deuterated (d 5 -) 1-phenyl-3-methyl-5-pyrazolone (PMP). The obtained O-glycan derivatives feature a permanent 10-Da mass difference between the d 0 - and d 5 -PMP forms, allowing complete discrimination and comparative quantification of these isotopically labeled O-glycans by mass spectrometric techniques. Moreover, the d 0 - and d 5 -PMP derivatives of O-glycans also have a relatively high hydrophobicity as well as a strong UV adsorption, especially suitable for high-resolution separation and high-sensitivity detection by RP-HPLC-UV. We have refined the conditions for the one-pot reaction as well as the corresponding sample purification approach. The good quantitation feasibility, reliability and linearity of this strategy have been verified using bovine fetuin and porcine stomach mucin as model O-glycoproteins. Additionally, we have also successfully applied this method to the quantitative

  13. Stable isotope labeling of glycoprotein expressed in silkworms using immunoglobulin G as a test molecule

    Energy Technology Data Exchange (ETDEWEB)

    Yagi, Hirokazu [Nagoya City University, Faculty and Graduate School of Pharmaceutical Sciences (Japan); Nakamura, Masatoshi [National Institute of Agrobiological Sciences, Genetic Resources Conservation Research Unit, Genetic Resources Center (Japan); Yokoyama, Jun [Taiyo Nippon Sanso Corporation, Tsukuba Laboratories (Japan); Zhang, Ying; Yamaguchi, Takumi [National Institutes of Natural Sciences, Institute for Molecular Science and Okazaki Institute for Integrative Bioscience (Japan); Kondo, Sachiko [Nagoya City University, Faculty and Graduate School of Pharmaceutical Sciences (Japan); Kobayashi, Jun [Yamaguchi University, Department of Biological and Environmental Sciences, Faculty of Agriculture (Japan); Kato, Tatsuya; Park, Enoch Y. [Shizuoka University, Laboratory of Biotechnology, Research Institute of Green Science and Technology (Japan); Nakazawa, Shiori [Nagoya University, Sugashima Marine Biological Laboratory, Graduate School of Science (Japan); Hashii, Noritaka; Kawasaki, Nana [National Institute of Health Sciences, Division of Biological Chemistry and Biologicals (Japan); Kato, Koichi, E-mail: kkato@phar.nagoya-cu.ac.jp [Nagoya City University, Faculty and Graduate School of Pharmaceutical Sciences (Japan)

    2015-06-15

    Silkworms serve as promising bioreactors for the production of recombinant proteins, including glycoproteins and membrane proteins, for structural and functional protein analyses. However, lack of methodology for stable isotope labeling has been a major deterrent to using this expression system for nuclear magnetic resonance (NMR) structural biology. Here we developed a metabolic isotope labeling technique using commercially available silkworm larvae. The fifth instar larvae were infected with baculoviruses for co-expression of recombinant human immunoglobulin G (IgG) as a test molecule, with calnexin as a chaperone. They were subsequently reared on an artificial diet containing {sup 15}N-labeled yeast crude protein extract. We harvested 0.1 mg of IgG from larva with a {sup 15}N-enrichment ratio of approximately 80 %. This allowed us to compare NMR spectral data of the Fc fragment cleaved from the silkworm-produced IgG with those of an authentic Fc glycoprotein derived from mammalian cells. Therefore, we successfully demonstrated that our method enables production of isotopically labeled glycoproteins for NMR studies.

  14. UNiquant, a Program for Quantitative Proteomics Analysis Using Stable Isotope Labeling

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Xin; Tolmachev, Aleksey V.; Shen, Yulei; Liu, Miao; Huang, Lin; Zhang, Zhixin; Anderson, Gordon A.; Smith, Richard D.; Chan, Wing C.; Hinrichs, Steven; Fu, Kai; Ding, Shi-Jian

    2011-03-04

    We present UNiquant, a new software program for analyzing stable isotope labeling (SIL) based quantitative proteomics data. UNiquant surpassed the performance of two other platforms, MaxQuant and Mascot Distiller, using complex proteome mixtures having either known or unknown heavy/light ratios. UNiquant is compatible with a broad spectrum of search engines and SIL methods, providing outstanding peptide pair identification and accurate measurement of the relative peptide/protein abundance.

  15. The synthesis of a tritium, carbon-14, and stable isotope-labeled cathepsin C inhibitors.

    Science.gov (United States)

    Allen, Paul; Bragg, Ryan A; Caffrey, Moya; Ericsson, Cecilia; Hickey, Michael J; Kingston, Lee P; Elmore, Charles S

    2017-02-01

    As part of a medicinal chemistry program aimed at developing a highly potent and selective cathepsin C inhibitor, tritium, carbon-14, and stable isotope-labeled materials were required. The synthesis of tritium-labeled methanesulfonate 5 was achieved via catalytic tritiolysis of a chloro precursor, albeit at a low radiochemical purity of 67%. Tritium-labeled AZD5248 was prepared via a 3-stage synthesis, utilizing amide-directed hydrogen isotope exchange. Carbon-14 and stable isotope-labeled AZD5248 were successfully prepared through modifications of the medicinal chemistry synthetic route, enabling the use of available labeled intermediates. Copyright © 2016 John Wiley & Sons, Ltd.

  16. Development of versatile isotopic labeling reagents for profiling the amine submetabolome by liquid chromatography–mass spectrometry

    International Nuclear Information System (INIS)

    Zhou, Ruokun; Huan, Tao; Li, Liang

    2015-01-01

    Highlights: • Two new reagents were developed for chemical isotope labeling mass spectrometry (MS). • They could be used to label amine-containing metabolites in a metabolomic sample. • The labeled metabolites could be detected with much improved sensitivity in MS. • One of the reagents could also help generate useful MS/MS spectra for structural analysis. • These reagents should be useful for quantitative metabolomics. - Abstract: Metabolomic profiling involves relative quantification of metabolites in comparative samples and identification of the significant metabolites that differentiate different groups (e.g., diseased vs. controls). Chemical isotope labeling (CIL) liquid chromatography–mass spectrometry (LC–MS) is an enabling technique that can provide improved metabolome coverage and metabolite quantification. However, chemical identification of labeled metabolites can still be a challenge. In this work, a new set of isotopic labeling reagents offering versatile properties to enhance both detection and identification are described. They were prepared by a glycine molecule (or its isotopic counterpart) and an aromatic acid with varying structures through a simple three-step synthesis route. In addition to relatively low costs of synthesizing the reagents, this reaction route allows adjusting reagent property in accordance with the desired application objective. To date, two isotopic reagents, 4-dimethylaminobenzoylamido acetic acid N-hydroxylsuccinimide ester (DBAA-NHS) and 4-methoxybenzoylamido acetic acid N-hydroxylsuccinimide ester (MBAA-NHS), for labeling the amine-containing metabolites (i.e., amine submetabolome) have been synthesized. The labeling conditions and the related LC–MS method have been optimized. We demonstrate that DBAA labeling can increase the metabolite detectability because of the presence of an electrospray ionization (ESI)-active dimethylaminobenzoyl group. On the other hand, MBAA labeled metabolites can be fragmented

  17. Development of versatile isotopic labeling reagents for profiling the amine submetabolome by liquid chromatography–mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Ruokun; Huan, Tao; Li, Liang, E-mail: Liang.Li@ualberta.ca

    2015-06-30

    Highlights: • Two new reagents were developed for chemical isotope labeling mass spectrometry (MS). • They could be used to label amine-containing metabolites in a metabolomic sample. • The labeled metabolites could be detected with much improved sensitivity in MS. • One of the reagents could also help generate useful MS/MS spectra for structural analysis. • These reagents should be useful for quantitative metabolomics. - Abstract: Metabolomic profiling involves relative quantification of metabolites in comparative samples and identification of the significant metabolites that differentiate different groups (e.g., diseased vs. controls). Chemical isotope labeling (CIL) liquid chromatography–mass spectrometry (LC–MS) is an enabling technique that can provide improved metabolome coverage and metabolite quantification. However, chemical identification of labeled metabolites can still be a challenge. In this work, a new set of isotopic labeling reagents offering versatile properties to enhance both detection and identification are described. They were prepared by a glycine molecule (or its isotopic counterpart) and an aromatic acid with varying structures through a simple three-step synthesis route. In addition to relatively low costs of synthesizing the reagents, this reaction route allows adjusting reagent property in accordance with the desired application objective. To date, two isotopic reagents, 4-dimethylaminobenzoylamido acetic acid N-hydroxylsuccinimide ester (DBAA-NHS) and 4-methoxybenzoylamido acetic acid N-hydroxylsuccinimide ester (MBAA-NHS), for labeling the amine-containing metabolites (i.e., amine submetabolome) have been synthesized. The labeling conditions and the related LC–MS method have been optimized. We demonstrate that DBAA labeling can increase the metabolite detectability because of the presence of an electrospray ionization (ESI)-active dimethylaminobenzoyl group. On the other hand, MBAA labeled metabolites can be fragmented

  18. Quantification of isotope-labelled and unlabelled folates in plasma, ileostomy and food samples.

    Science.gov (United States)

    Büttner, Barbara E; Öhrvik, Veronica E; Witthöft, Cornelia M; Rychlik, Michael

    2011-01-01

    New stable isotope dilution assays were developed for the simultaneous quantitation of [(13)C(5)]-labelled and unlabelled 5-methyltetrahydrofolic acid, 5-formyltetrahydrofolic acid, folic acid along with unlabelled tetrahydrofolic acid and 10-formylfolic acid in clinical samples deriving from human bioavailability studies, i.e. plasma, ileostomy samples, and food. The methods were based on clean-up by strong anion exchange followed by LC-MS/MS detection. Deuterated analogues of the folates were applied as the internal standards in the stable isotope dilution assays. Assay sensitivity was sufficient to detect all relevant folates in the respective samples as their limits of detection were below 0.62 nmol/L in plasma and below 0.73 μg/100 g in food or ileostomy samples. Quantification of the [(13)C(5)]-label in clinical samples offers the possibility to differentiate between folate from endogenous body pools and the administered dose when executing bioavailability trials.

  19. Method and device for isotope separation

    International Nuclear Information System (INIS)

    Dawson, J.M.

    1976-01-01

    The method works with a converted Q machine. The plasma containing the isotopes to be separated is crossed by a magnetic field running in the direction of the plasma column. More energy is transfered to the chosen isotope by oscillating magnetic and/or electric fields or by sound waves by using the specific resonance frequency for the selected isotope. The isotopes thus heated to different extents can be separated according to various methods given in the patent claims. (GG) [de

  20. Isotopic Abundance and Chemical Purity Analysis of Stable Isotope Deuterium Labeled Sudan I

    Directory of Open Access Journals (Sweden)

    CAI Yin-ping;LEI Wen;ZHENG Bo;DU Xiao-ning

    2014-02-01

    Full Text Available It is important that to analysis of the isotopic abundance and chemical purity of Sudan I-D5, which is the internal standard of isotope dilution mass spectrometry. The isotopic abundance of Sudan I-D5 is detected by “mass cluster” classification method and LC-MS. The repeatability and reproducibility experiments were carried out by using different mass spectrometers and different operators. The RSD was less than 0.1%, so the repeatability and reproducibility were satisfactory. The accuracy and precision of the isotopic abundance analysis method was good with the results of F test and t test. The high performance liquid chromatography (HPLC had been used for detecting the chemical purity of Sudan I-D5 as external standard method.

  1. METHOD AND MODULE FOR OPTICAL SUBCARRIER LABELLING

    DEFF Research Database (Denmark)

    2004-01-01

    The present invention relates to optical labelling in WDM networks, in that it provides a method and a module to be used in subcarrier label generation and switching in network edge nodes and core switch nodes. The methods and modules are typically employed in Optical Subcarrier Multiplexing (OSCM......) transmitters. The payload and the label are encoded independently on optical carrier and subcarrier signals respectively, using electro-optical modulators. The invention applies single or double sideband carrier-suppressed modulation to generate subcarrier signals for encoding of the label. Thereby the payload...... encoded carrier signal and the label encoded subcarrier signal can be coupled directly without prior filtering....

  2. Efficient segmental isotope labeling of multi-domain proteins using Sortase A

    Energy Technology Data Exchange (ETDEWEB)

    Freiburger, Lee, E-mail: lee.freiburger@tum.de; Sonntag, Miriam, E-mail: miriam.sonntag@mytum.de; Hennig, Janosch, E-mail: janosch.hennig@helmholtz-muenchen.de [Helmholtz Zentrum München, Institute of Structural Biology (Germany); Li, Jian, E-mail: lijianzhongbei@163.com [Chinese Academy of Sciences, Tianjin Institute of Industrial Biotechnology (China); Zou, Peijian, E-mail: peijian.zou@helmholtz-muenchen.de; Sattler, Michael, E-mail: sattler@helmholtz-muenchen.de [Helmholtz Zentrum München, Institute of Structural Biology (Germany)

    2015-09-15

    NMR studies of multi-domain protein complexes provide unique insight into their molecular interactions and dynamics in solution. For large proteins domain-selective isotope labeling is desired to reduce signal overlap, but available methods require extensive optimization and often give poor ligation yields. We present an optimized strategy for segmental labeling of multi-domain proteins using the S. aureus transpeptidase Sortase A. Critical improvements compared to existing protocols are (1) the efficient removal of cleaved peptide fragments by centrifugal filtration and (2) a strategic design of cleavable and non-cleavable affinity tags for purification. Our approach enables routine production of milligram amounts of purified segmentally labeled protein for NMR and other biophysical studies.

  3. Sequencing of Isotope-Labeled Small RNA Using Femtosecond Laser Ablation Time-of-Flight Mass Spectrometry

    Science.gov (United States)

    Kurata-Nishimura, Mizuki; Ando, Yoshinari; Kobayashi, Tohru; Matsuo, Yukari; Suzuki, Harukazu; Hayashizaki, Yoshihide; Kawai, Jun

    2010-04-01

    A novel method for the analysis of sequences of small RNAs using nucleotide triphosphates labeled with stable isotopes has been developed using time-of-flight mass spectroscopy combined with femtosecond laser ablation (fsLA-TOF-MS). Small RNAs synthesized with nucleotides enriched in 13C and 15N were efficiently atomized and ionized by single-shot fsLA and the isotope ratios 13C/12C and 15N/14N were evaluated using the TOF-MS method. By comparing the isotope ratios among four different configurations, the number of nucleotide contents of the control RNA sample were successfully reproduced.

  4. Identifying and quantifying proteolytic events and the natural N terminome by terminal amine isotopic labeling of substrates.

    Science.gov (United States)

    Kleifeld, Oded; Doucet, Alain; Prudova, Anna; auf dem Keller, Ulrich; Gioia, Magda; Kizhakkedathu, Jayachandran N; Overall, Christopher M

    2011-09-22

    Analysis of the sequence and nature of protein N termini has many applications. Defining the termini of proteins for proteome annotation in the Human Proteome Project is of increasing importance. Terminomics analysis of protease cleavage sites in degradomics for substrate discovery is a key new application. Here we describe the step-by-step procedures for performing terminal amine isotopic labeling of substrates (TAILS), a 2- to 3-d (depending on method of labeling) high-throughput method to identify and distinguish protease-generated neo-N termini from mature protein N termini with all natural modifications with high confidence. TAILS uses negative selection to enrich for all N-terminal peptides and uses primary amine labeling-based quantification as the discriminating factor. Labeling is versatile and suited to many applications, including biochemical and cell culture analyses in vitro; in vivo analyses using tissue samples from animal and human sources can also be readily performed. At the protein level, N-terminal and lysine amines are blocked by dimethylation (formaldehyde/sodium cyanoborohydride) and isotopically labeled by incorporating heavy and light dimethylation reagents or stable isotope labeling with amino acids in cell culture labels. Alternatively, easy multiplex sample analysis can be achieved using amine blocking and labeling with isobaric tags for relative and absolute quantification, also known as iTRAQ. After tryptic digestion, N-terminal peptide separation is achieved using a high-molecular-weight dendritic polyglycerol aldehyde polymer that binds internal tryptic and C-terminal peptides that now have N-terminal alpha amines. The unbound naturally blocked (acetylation, cyclization, methylation and so on) or labeled mature N-terminal and neo-N-terminal peptides are recovered by ultrafiltration and analyzed by tandem mass spectrometry (MS/MS). Hierarchical substrate winnowing discriminates substrates from the background proteolysis products and

  5. Method for separating gaseous mixtures of isotopes

    International Nuclear Information System (INIS)

    Neimann, H.J.; Schuster, E.; Kersting, A.

    1976-01-01

    A gaseous mixture of isotopes is separated by laser excitation of the isotope mixture with a narrow band of wavelengths, molecularly exciting mainly the isotope to be separated and thereby promoting its reaction with its chemical partner which is excited in a separate chamber. The excited isotopes and the chemical partner are mixed, perhaps in a reaction chamber to which the two excited components are conducted by very short conduits. The improvement of this method is the physical separation of the isotope mixture and its partner during excitation. The reaction between HCl and the mixture of 238 UF 6 and 235 UF 6 is discussed

  6. Extrinsic labelling of staple food crops with isotopic iron does not consistently result in full equilibration: Revisiting the methodology

    Science.gov (United States)

    Extrinsic isotopic labeling of food Fe has been used for over 50 years to measure Fe absorption. This method is based on the assumption that complete equilibration occurs between the extrinsic and the intrinsic Fe prior to intestinal absorption. The present study tested this assumption via use of in...

  7. NMR studies of two spliced leader RNAs using isotope labeling

    Energy Technology Data Exchange (ETDEWEB)

    Lapham, J.; Crothers, D.M. [Yale Univ., New Haven, CT (United States)

    1994-12-01

    Spliced leader RNAs are a class of RNA molecules (<200 nts) involved in the trans splicing of messenger RNA found in trypanosomes, nematodes, and other lower eukaryotes. The spliced leader RNA from the trypanosome Leptomonas Collosoma exists in two alternate structural forms with similar thermal stabilities. The 54 nucleotides on the 5{prime} end of the SL molecule is structurally independent from the 3{prime} half of the RNA, and displays the two structural forms. Furthermore, the favored of the two structures was shown to contain anomalous nuclease sensitivity and thermal stability features, which suggests that there may be tertiary interactions between the splice site and other nucleotides in the 5{prime} end. Multidimensional NMR studies are underway to elucidate the structural elements present in the SL RNAs that give rise to their physical properties. Two spliced leader sequences have been studied. The first, the 54 nucleotides on the 5{prime} end of the L. Collosoma sequence, was selected because of earlier studies in our laboratory. The second sequence is the 5{prime} end of the trypanosome Crithidia Fasciculata, which was chosen because of its greater sequence homology to other SL sequences. Given the complexity of the NMR spectra for RNA molecules of this size, we have incorporated {sup 15}N/{sup 13}C-labeled nucleotides into the RNA. One of the techniques we have developed to simplify the spectra of these RNA molecules is isotope labeling of specific regions of the RNA. This has been especially helpful in assigning the secondary structure of molecules that may be able to adopt multiple conformations. Using this technique one can examine a part of the molecule without spectral interference from the unlabeled portion. We hope this approach will promote an avenue for studying the structure of larger RNAs in their native surroundings.

  8. Preparation of H3-labelled methyl ethers of saturated fatty acids by heterogeneous catalytic isotope exchange in solution with gaseous tritium

    International Nuclear Information System (INIS)

    Shevchenko, V.P.; Myasoedov, N.F.

    1980-01-01

    A simple method of preparing 3 H-labelled methyl ethers of saturated fatty acids in the dioxane solution using the method of isotopic heterogenous catalytic exchange with gaseous tritium, is suggested. 3 H-labelled natural fatty acids (C 12 -C 18 ) are prepared by alkaline hydrolysis [ru

  9. Dansylation isotope labeling liquid chromatography mass spectrometry for parallel profiling of human urinary and fecal submetabolomes

    Energy Technology Data Exchange (ETDEWEB)

    Su, Xiaoling [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Wang, Nan [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2 (Canada); Chen, Deying [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Li, Yunong [Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2 (Canada); Lu, Yingfeng [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Huan, Tao [Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2 (Canada); Xu, Wei [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Li, Liang, E-mail: Liang.Li@ualberta.ca [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2 (Canada); Li, Lanjuan, E-mail: ljli@zju.edu.cn [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China)

    2016-01-15

    Human urine and feces can be non-invasively collected for metabolomics-based disease biomarker discovery research. Because urinary and fecal metabolomes are thought to be different, analysis of both biospecimens may generate a more comprehensive metabolomic profile that can be better related to the health state of an individual. Herein we describe a method of using differential chemical isotope labeling (CIL) liquid chromatography mass spectrometry (LC-MS) for parallel metabolomic profiling of urine and feces. Dansylation labeling was used to quantify the amine/phenol submetabolome changes among different samples based on {sup 12}C-labeling of individual samples and {sup 13}C-labeling of a pooled urine or pooled feces and subsequent analysis of the {sup 13}C-/{sup 12}C-labeled mixture by LC-MS. The pooled urine and pooled feces are further differentially labeled, mixed and then analyzed by LC-MS in order to relate the metabolite concentrations of the common metabolites found in both biospecimens. This method offers a means of direct comparison of urinary and fecal submetabolomes. We evaluated the analytical performance and demonstrated the utility of this method in the analysis of urine and feces collected daily from three healthy individuals for 7 days. On average, 2534 ± 113 (n = 126) peak pairs or metabolites could be detected from a urine sample, while 2507 ± 77 (n = 63) peak pairs were detected from a fecal sample. In total, 5372 unique peak pairs were detected from all the samples combined; 3089 and 3012 pairs were found in urine and feces, respectively. These results reveal that the urine and fecal metabolomes are very different, thereby justifying the consideration of using both biospecimens to increase the probability of finding specific biomarkers of diseases. Furthermore, the CIL LC-MS method described can be used to perform parallel quantitative analysis of urine and feces, resulting in more complete coverage of the human metabolome

  10. Dansylation isotope labeling liquid chromatography mass spectrometry for parallel profiling of human urinary and fecal submetabolomes

    International Nuclear Information System (INIS)

    Su, Xiaoling; Wang, Nan; Chen, Deying; Li, Yunong; Lu, Yingfeng; Huan, Tao; Xu, Wei; Li, Liang; Li, Lanjuan

    2016-01-01

    Human urine and feces can be non-invasively collected for metabolomics-based disease biomarker discovery research. Because urinary and fecal metabolomes are thought to be different, analysis of both biospecimens may generate a more comprehensive metabolomic profile that can be better related to the health state of an individual. Herein we describe a method of using differential chemical isotope labeling (CIL) liquid chromatography mass spectrometry (LC-MS) for parallel metabolomic profiling of urine and feces. Dansylation labeling was used to quantify the amine/phenol submetabolome changes among different samples based on "1"2C-labeling of individual samples and "1"3C-labeling of a pooled urine or pooled feces and subsequent analysis of the "1"3C-/"1"2C-labeled mixture by LC-MS. The pooled urine and pooled feces are further differentially labeled, mixed and then analyzed by LC-MS in order to relate the metabolite concentrations of the common metabolites found in both biospecimens. This method offers a means of direct comparison of urinary and fecal submetabolomes. We evaluated the analytical performance and demonstrated the utility of this method in the analysis of urine and feces collected daily from three healthy individuals for 7 days. On average, 2534 ± 113 (n = 126) peak pairs or metabolites could be detected from a urine sample, while 2507 ± 77 (n = 63) peak pairs were detected from a fecal sample. In total, 5372 unique peak pairs were detected from all the samples combined; 3089 and 3012 pairs were found in urine and feces, respectively. These results reveal that the urine and fecal metabolomes are very different, thereby justifying the consideration of using both biospecimens to increase the probability of finding specific biomarkers of diseases. Furthermore, the CIL LC-MS method described can be used to perform parallel quantitative analysis of urine and feces, resulting in more complete coverage of the human metabolome. - Highlights: • A

  11. Pyrolysis of Cigarette Ingredients Labelled with Stable Isotopes

    Directory of Open Access Journals (Sweden)

    Stotesbury S

    2014-12-01

    Full Text Available It is important to know how tobacco additives behave when cigarettes are smoked, whether they transfer intact to the smoke or whether there is any decomposition during smoking. Pyrolysis-GC-MS is a technique that can be focussed upon the effects of combustion from a single material free from interference from the complex mixture of different components present in the smoke. However, because pyrolysis is a model technique, the results need to be validated by comparison with cigarette smoke chemistry. In a previous paper we presented such a method for modelling the smoke chemistry from a burning cigarette using pyrolysis-GC-MS. The transfer and the extent of degradation of anisole, p-anisaldehyde, benzaldehyde, isoamylisovalerate, methyl trans-cinnamate and vanillin within a burning cigarette were estimated using this pyrolysis method. When these data were compared with results from smoke studies from 14C-analogues of the materials, the high levels of transfer predicted by pyrolysis were found to be generally consistent with the smoke chemistry data. However, there were still two outstanding issues. Firstly, there was some ambiguity in the labelled study about whether vanillin actually transferred without degradation or not. Furthermore, the results from the 14C-labelled study showed a greater extent of degradation for p-anisaldehyde than that indicated from the pyrolysis experiments. The purpose of the current study was to present some new information obtained to address these questions by better understanding the effect upon the smoke chemistry from adding vanillin and p-anisaldehyde, and the relationship between the smoke chemistry and the pyrolysis results. Components were identified in the smoke from cigarettes loaded with p-anisaldehyde and vanillin labelled with 18O and 13C. The extent of degradation from each additive was estimated by identifying labelled degradation products in the smoke. Because there was a clear distinction between the

  12. Isotopic labeling affects 1,25-dihydroxyvitamin D metabolism

    International Nuclear Information System (INIS)

    Halloran, B.P.; Bikle, D.D.; Castro, M.E.; Gee, E.

    1989-01-01

    Isotope substitution can change the biochemical properties of vitamin D. To determine the effect of substituting 3H for 1H on the metabolism of 1,25(OH)2D3, we measured the metabolic clearance rate and renal metabolism of unlabeled and 3H-labeled 1,25(OH)2D3. Substitution of 3H for 1H on carbons 26 and 27 [1,25(OH)2[26,27(n)-3H]D3] or on carbons 23 and 24 [1,25(OH)2[23,24(n)-3H]D3] reduced the in vivo metabolic clearance rate of 1,25(OH)2D3 by 36% and 37%, respectively, and reduced the in vitro renal catabolism of 1,25(OH)2D3 by 11% and 54%, respectively. Substitutions of 3H for 1H on carbons 23 and 24 as opposed to carbons 26 and 27 reduced conversion of [3H]1,25(OH)2D3 to [3H]1,24,25(OH)2D3 by 25% and to putative 24-oxo-1,23,25-dihydroxyvitamin D3 by 1600%. These results indicate that substitution of 3H for 1H on carbons 26 and 27 or on carbons 23 and 24 can reduce the metabolic clearance rate and in vitro metabolism of 1,25(OH)2D3 and quantitatively alter the pattern of metabolic products produced

  13. Production of large quantities of isotopically labeled protein in Pichia pastoris by fermentation

    International Nuclear Information System (INIS)

    Wood, Matthew J.; Komives, Elizabeth A.

    1999-01-01

    Heterologous expression in Pichia pastoris has many of the advantages of eukaryotic expression, proper folding and disulfide bond formation, glycosylation, and secretion. Contrary to other eukaryotic systems, protein production from P.pastoris occurs in simple minimal defined media making this system attractive for production of labeled proteins for NMR analysis. P.pastoris is therefore the expression system of choice for NMR of proteins that cannot be refolded from inclusion bodies or that require post-translational modifications for proper folding or function. The yield of expressed proteins from P.pastoris depends critically on growth conditions, and attainment of high cell densities by fermentation has been shown to improve protein yields by 10-100-fold. Unfortunately, the cost of the isotopically enriched fermentation media components, particularly 15NH4OH, is prohibitively high. We report fermentation methods that allow for both 15N- labeling from (15NH4)2SO4 and 13C-labeling from 13C-glucose or 13C-glycerol of proteins produced in Pichia pastoris. Expression of an 83 amino acid fragment of thrombomodulin with two N-linked glycosylation sites shows that fermentation is more cost effective than shake flask growth for isotopic enrichment

  14. Stable isotope labelling with amino acids in cell culture for human embryonic stem cell proteomic analysis

    DEFF Research Database (Denmark)

    Harkness, Linda; Prokhorova, Tatyana A; Kassem, Moustapha

    2012-01-01

    The identification and quantitative measurements of proteins in human embryonic stem cells (hESC) is a fast growing interdisciplinary area with an enormous impact on understanding the biology of hESC and the mechanism controlling self-renewal and differentiation. Using a quantitative mass...... spectroscopic method of stable isotope labelling with amino acids during cell culture (SILAC), we are able to analyse differential expression of proteins from different cellular compartments and to identify intracellular signalling pathways involved in self-renewal and differentiation. In this chapter, we...

  15. Method for isotope separation by photodeflection

    International Nuclear Information System (INIS)

    Bernhardt, A.F.

    1977-01-01

    In the method of separating isotopes wherein a desired isotope species is selectively deflected out of a beam of mixed isotopes by irradiating the beam with a directed beam of light of narrowly defined frequency which is selectively absorbed by the desired species, the improvement comprising irradiating the deflected beam with light from other light sources whose frequencies are selected to cause the depopulation of any metastable excited states

  16. Improved segmental isotope labeling of proteins and application to a larger protein

    International Nuclear Information System (INIS)

    Otomo, Takanori; Teruya, Kenta; Uegaki, Koichi; Yamazaki, Toshio; Kyogoku, Yoshimasa

    1999-01-01

    A new isotope labeling technique for peptide segments in a protein sample was recently established using the protein splicing element intein [Yamazaki et al. (1998) J. Am. Chem. Soc., 120, 5591-5592]. This method makes it possible to observe signals of a selected amino (N-) or carboxyl (C-) terminal region along a peptide chain. However, there is a problem with the yield of the segmentally labeled protein. In this paper, we report an increase in the yield of the protein that enables the production of sufficient amounts of segmentally 13 C/ 15 N-labeled protein samples. This was achieved by improvement of the expression level of the N-terminal fragment in cells and the efficiency of refolding into the active splicing conformation. The N-terminal fragment was expressed as a fused protein with the cellulose binding domain at its N-terminus, which was expressed as an insoluble peptide in cells and the expression level was increased. Incubation with 2.5 M urea and 50% glycerol increased the efficiency of the refolding greatly, thereby raising the final yields of the ligated proteins. The feasibility of application of the method to a high-molecular-weight protein was demonstrated by the results for a maltose binding protein consisting of 370 amino acids. All four examined joints in the maltose binding protein were successfully ligated to produce segmentally labeled protein samples

  17. Multiple tag labeling method for DNA sequencing

    Science.gov (United States)

    Mathies, R.A.; Huang, X.C.; Quesada, M.A.

    1995-07-25

    A DNA sequencing method is described which uses single lane or channel electrophoresis. Sequencing fragments are separated in the lane and detected using a laser-excited, confocal fluorescence scanner. Each set of DNA sequencing fragments is separated in the same lane and then distinguished using a binary coding scheme employing only two different fluorescent labels. Also described is a method of using radioisotope labels. 5 figs.

  18. Noninvasive imaging of protein metabolic labeling in single human cells using stable isotopes and Raman microscopy

    NARCIS (Netherlands)

    van Manen, H.J.; Lenferink, Aufrid T.M.; Otto, Cornelis

    2008-01-01

    We have combined nonresonant Raman microspectroscopy and spectral imaging with stable isotope labeling by amino acids in cell culture (SILAC) to selectively detect the incorporation of deuterium-labeled phenylalanine, tyrosine, and methionine into proteins in intact, single HeLa cells. The C−D

  19. Synthesis of {sup 15}N isotope labeled alanine; Sintese da alanina enriquecida com {sup 15}N

    Energy Technology Data Exchange (ETDEWEB)

    Oliveira, Claudineia R. de; Bendassolli, Jose Albertino; Sant' Ana, Carlos Roberto; Tagliassachi, Romulo Barbieri; Maximo, Everaldo; Prestes, Clelber Vieira [Centro de Energia Nuclear na Agricultura (CENA), Piracicaba, SP (Brazil). Dept. de Isotopos Estaveis]. E-mail: crolivei@cena.usp.br

    2005-07-01

    The application of light chemical elements and their stable isotopes in biological studies have been increased over the last years. The use of {sup 15}N labeled amino acids is an important tool for elucidation of peptides structures. This paper describe a method for the synthesis of {sup 15}N isotope labeled alanine at lower costs than international ones, as well as the details of the recovery system of the nitrogen residues. In the present work an amination of {alpha}-haloacids, with the bromopropionic carboxylic acid and labeled aqua ammonia ({sup 15}NH{sub 3} aq) was carried out. In order to avoid eventually losses of {sup 15}NH{sub 3}, special cares were adopted, since the production cost is high. Although the acquisition cost of the {sup 13}N (radioactive) labeled compounds is lower, the obtained stable tracer will allow the accomplishment of important studies of the nitrogen cycling in living things, less occupational and environment hazards, and the time limitation problems in field studies. The tests took place in triplicates with NH{sub 3} (aq) being employed. With the establishment of the system for {sup 15}NH{sub 3} recovery, an average of 94 % of the ammonia employed in the synthesis process was recovered. The purity of the amino acid was state determined by TLC (Thin Layer Chromatography) and HPLC (High-Performance Liquid Chromatography) with a fluorescence detector. The Rf and the retention time of the synthesized sample were similar the sigma standard. Finally, regarding the established conditions, it was possible to obtain the alanine with a production cost about 40 % lower than the international price. (author)

  20. Evaluation of three methods of platelet labelling

    International Nuclear Information System (INIS)

    Mortelmans, L.; Verbruggen, A.; Roo, M. de; Vermylen, J.

    1986-01-01

    The study of the kinetics of labelled platelets makes sense only when the platelets preserve their viability after separation and labelling. The separation and labelling procedures described in the manual of two producers of 111 In-oxinate (Amersham, Mallinckrodt) have been evaluated by in vitro aggregation tests. The method of Mallinckrodt diminished the aggregation capacities of the thrombocytes. The labelled platelets with normal in vitro aggregation response (Amersham) were tested in vivo in 11 patients who underwent peripheral bypass surgery. The platelet half-life and the platelet accumulation on bypass grafts were checked one week post-operatively. Because of the poor in vivo response of both methods (exponential half-life curve and bad graft visualization), a third method based on that described by W.A. Heaton et al. 1979 was optimized in the authors' laboratory with good in vitro and in vivo results in 12 patients. (author)

  1. Evaluation of three methods of platelet labelling.

    Science.gov (United States)

    Mortelmans, L; Verbruggen, A; De Roo, M; Vermylen, J

    1986-07-01

    The study of the kinetics of labelled platelets makes sense only when the platelets preserve their viability after separation and labelling. The separation and labelling procedures described in the manual of two producers of 111In-oxinate (Amersham, Mallinckrodt) have been evaluated by in vitro aggregation tests. The method of Mallinckrodt diminished the aggregation capacities of the thrombocytes. The labelled platelets with normal in vitro aggregation response (Amersham) were tested in vivo in 11 patients who underwent peripheral bypass surgery. The platelet half-life and the platelet accumulation on bypass grafts were checked one week post-operatively. Because of the poor in vivo response of both methods (exponential half-life curve and bad graft visualization), a third method was optimized in our laboratory with good in vitro and in vivo results in 12 patients.

  2. Isotopic labelling studies for a gold-catalysed skeletal rearrangement of alkynyl aziridines

    Directory of Open Access Journals (Sweden)

    Neil Spencer

    2011-06-01

    Full Text Available Isotopic labelling studies were performed to probe a proposed 1,2-aryl shift in the gold-catalysed cycloisomerisation of alkynyl aziridines into 2,4-disubstituted pyrroles. Two isotopomers of the expected skeletal rearrangement product were identified using 13C-labelling and led to a revised mechanism featuring two distinct skeletal rearrangements. The mechanistic proposal has been rationalised against the reaction of a range of 13C- and deuterium-labelled substrates.

  3. Highly efficient residue-selective labeling with isotope-labeled Ile, Leu, and Val using a new auxotrophic E. coli strain

    International Nuclear Information System (INIS)

    Miyanoiri, Yohei; Ishida, Yojiro; Takeda, Mitsuhiro; Terauchi, Tsutomu; Inouye, Masayori; Kainosho, Masatsune

    2016-01-01

    We recently developed a practical protocol for preparing proteins bearing stereo-selectively 13 C-methyl labeled leucines and valines, instead of the commonly used 13 C-methyl labeled precursors for these amino acids, by E. coli cellular expression. Using this protocol, proteins with any combinations of isotope-labeled or unlabeled Leu and Val residues were prepared, including some that could not be prepared by the precursor methods. However, there is still room for improvement in the labeling efficiencies for Val residues, using the methods with labeled precursors or Val itself. This is due to the fact that the biosynthesis of Val could not be sufficiently suppressed, even by the addition of large amounts of Val or its precursors. In this study, we completely solved this problem by using a mutant strain derived from E. coli BL21(DE3), in which the metabolic pathways depending on two enzymes, dihydroxy acid dehydratase and β-isopropylmalate dehydrogenase, are completely aborted by deleting the ilvD and leuB genes, which respectively encode these enzymes. The ΔilvD E. coli mutant terminates the conversion from α,β-dihydroxyisovalerate to α-ketoisovalerate, and the conversion from α,β-dihydroxy-α-methylvalerate to α-keto-β-methylvalerate, which produce the preceding precursors for Val and Ile, respectively. By the further deletion of the leuB gene, the conversion from Val to Leu was also fully terminated. Taking advantage of the double-deletion mutant, ΔilvDΔleuB E. coli BL21(DE3), an efficient and residue-selective labeling method with various isotope-labeled Ile, Leu, and Val residues was established.

  4. Highly efficient residue-selective labeling with isotope-labeled Ile, Leu, and Val using a new auxotrophic E. coli strain

    Energy Technology Data Exchange (ETDEWEB)

    Miyanoiri, Yohei [Nagoya University, Structural Biology Research Center, Graduate School of Science (Japan); Ishida, Yojiro [Rutgers University-Robert Wood Johnson Medical School, Center for Advanced Biotechnology and Medicine (United States); Takeda, Mitsuhiro [Nagoya University, Structural Biology Research Center, Graduate School of Science (Japan); Terauchi, Tsutomu [Tokyo Metropolitan University, Graduate School of Science and Engineering (Japan); Inouye, Masayori [Rutgers University-Robert Wood Johnson Medical School, Center for Advanced Biotechnology and Medicine (United States); Kainosho, Masatsune, E-mail: kainosho@tmu.ac.jp [Nagoya University, Structural Biology Research Center, Graduate School of Science (Japan)

    2016-06-15

    We recently developed a practical protocol for preparing proteins bearing stereo-selectively {sup 13}C-methyl labeled leucines and valines, instead of the commonly used {sup 13}C-methyl labeled precursors for these amino acids, by E. coli cellular expression. Using this protocol, proteins with any combinations of isotope-labeled or unlabeled Leu and Val residues were prepared, including some that could not be prepared by the precursor methods. However, there is still room for improvement in the labeling efficiencies for Val residues, using the methods with labeled precursors or Val itself. This is due to the fact that the biosynthesis of Val could not be sufficiently suppressed, even by the addition of large amounts of Val or its precursors. In this study, we completely solved this problem by using a mutant strain derived from E. coli BL21(DE3), in which the metabolic pathways depending on two enzymes, dihydroxy acid dehydratase and β-isopropylmalate dehydrogenase, are completely aborted by deleting the ilvD and leuB genes, which respectively encode these enzymes. The ΔilvD E. coli mutant terminates the conversion from α,β-dihydroxyisovalerate to α-ketoisovalerate, and the conversion from α,β-dihydroxy-α-methylvalerate to α-keto-β-methylvalerate, which produce the preceding precursors for Val and Ile, respectively. By the further deletion of the leuB gene, the conversion from Val to Leu was also fully terminated. Taking advantage of the double-deletion mutant, ΔilvDΔleuB E. coli BL21(DE3), an efficient and residue-selective labeling method with various isotope-labeled Ile, Leu, and Val residues was established.

  5. Validation of single-sample doubly labeled water method

    International Nuclear Information System (INIS)

    Webster, M.D.; Weathers, W.W.

    1989-01-01

    We have experimentally validated a single-sample variant of the doubly labeled water method for measuring metabolic rate and water turnover in a very small passerine bird, the verdin (Auriparus flaviceps). We measured CO 2 production using the Haldane gravimetric technique and compared these values with estimates derived from isotopic data. Doubly labeled water results based on the one-sample calculations differed from Haldane values by less than 0.5% on average (range -8.3 to 11.2%, n = 9). Water flux computed by the single-sample method differed by -1.5% on average from results for the same birds based on the standard, two-sample technique (range -13.7 to 2.0%, n = 9)

  6. A method of uranium isotopes concentration analysis

    International Nuclear Information System (INIS)

    Lin Yuangen; Jiang Meng; Wu Changli; Duan Zhanyuan; Guo Chunying

    2010-01-01

    A basic method of uranium isotopes concentration is described in this paper. The iteration method is used to calculate the relative efficiency curve, by analyzing the characteristic γ energy spectrum of 235 U, 232 U and the daughter nuclide of 238 U, then the relative activity can be calculated, at last the uranium isotopes concentration can be worked out, and the result is validated by the experimentation. (authors)

  7. Efficient production of isotopically labeled proteins by cell-free synthesis: A practical protocol

    Energy Technology Data Exchange (ETDEWEB)

    Torizawa, Takuya; Shimizu, Masato [Crest, Jst (Japan); Taoka, Masato [Tokyo Metropolitan University, Graduate School of Science (Japan); Miyano, Hiroshi [Ajinomoto Co., Inc. Institute of Life Sciences (Japan); Kainosho, Masatsune [Crest, Jst (Japan)], E-mail: kainosho@nmr.chem.metro-u.ac.jp

    2004-11-15

    We provide detailed descriptions of our refined protocols for the cell-free production of labeled protein samples for NMR spectroscopy. These methods are efficient and overcome two critical problems associated with the use of conventional Escherichia coli extract systems. Endogenous amino acids normally present in E. coli S30 extracts dilute the added labeled amino acids and degrade the quality of NMR spectra of the target protein. This problem was solved by altering the protocol used in preparing the S30 extract so as to minimize the content of endogenous amino acids. The second problem encountered in conventional E. coli cell-free protein production is non-uniformity in the N-terminus of the target protein, which can complicate the NMR spectra. This problem was solved by adding a DNA sequence to the construct that codes for a cleavable N-terminal peptide tag. Addition of the tag serves to increase the yield of the protein as well as to ensure a homogeneous protein product following tag cleavage. We illustrate the method by describing its stepwise application to the production of calmodulin samples with different stable isotope labeling patterns for NMR analysis.

  8. Efficient production of isotopically labeled proteins by cell-free synthesis: A practical protocol

    International Nuclear Information System (INIS)

    Torizawa, Takuya; Shimizu, Masato; Taoka, Masato; Miyano, Hiroshi; Kainosho, Masatsune

    2004-01-01

    We provide detailed descriptions of our refined protocols for the cell-free production of labeled protein samples for NMR spectroscopy. These methods are efficient and overcome two critical problems associated with the use of conventional Escherichia coli extract systems. Endogenous amino acids normally present in E. coli S30 extracts dilute the added labeled amino acids and degrade the quality of NMR spectra of the target protein. This problem was solved by altering the protocol used in preparing the S30 extract so as to minimize the content of endogenous amino acids. The second problem encountered in conventional E. coli cell-free protein production is non-uniformity in the N-terminus of the target protein, which can complicate the NMR spectra. This problem was solved by adding a DNA sequence to the construct that codes for a cleavable N-terminal peptide tag. Addition of the tag serves to increase the yield of the protein as well as to ensure a homogeneous protein product following tag cleavage. We illustrate the method by describing its stepwise application to the production of calmodulin samples with different stable isotope labeling patterns for NMR analysis

  9. Isotope coded protein labeling coupled immunoprecipitation (ICPL-IP): a novel approach for quantitative protein complex analysis from native tissue.

    Science.gov (United States)

    Vogt, Andreas; Fuerholzner, Bettina; Kinkl, Norbert; Boldt, Karsten; Ueffing, Marius

    2013-05-01

    High confidence definition of protein interactions is an important objective toward the understanding of biological systems. Isotope labeling in combination with affinity-based isolation of protein complexes has increased in accuracy and reproducibility, yet, larger organisms--including humans--are hardly accessible to metabolic labeling and thus, a major limitation has been its restriction to small animals, cell lines, and yeast. As composition as well as the stoichiometry of protein complexes can significantly differ in primary tissues, there is a great demand for methods capable to combine the selectivity of affinity-based isolation as well as the accuracy and reproducibility of isotope-based labeling with its application toward analysis of protein interactions from intact tissue. Toward this goal, we combined isotope coded protein labeling (ICPL)(1) with immunoprecipitation (IP) and quantitative mass spectrometry (MS). ICPL-IP allows sensitive and accurate analysis of protein interactions from primary tissue. We applied ICPL-IP to immuno-isolate protein complexes from bovine retinal tissue. Protein complexes of immunoprecipitated β-tubulin, a highly abundant protein with known interactors as well as the lowly expressed small GTPase RhoA were analyzed. The results of both analyses demonstrate sensitive and selective identification of known as well as new protein interactions by our method.

  10. Isotope Coded Protein Labeling Coupled Immunoprecipitation (ICPL-IP): A Novel Approach for Quantitative Protein Complex Analysis From Native Tissue*

    Science.gov (United States)

    Vogt, Andreas; Fuerholzner, Bettina; Kinkl, Norbert; Boldt, Karsten; Ueffing, Marius

    2013-01-01

    High confidence definition of protein interactions is an important objective toward the understanding of biological systems. Isotope labeling in combination with affinity-based isolation of protein complexes has increased in accuracy and reproducibility, yet, larger organisms—including humans—are hardly accessible to metabolic labeling and thus, a major limitation has been its restriction to small animals, cell lines, and yeast. As composition as well as the stoichiometry of protein complexes can significantly differ in primary tissues, there is a great demand for methods capable to combine the selectivity of affinity-based isolation as well as the accuracy and reproducibility of isotope-based labeling with its application toward analysis of protein interactions from intact tissue. Toward this goal, we combined isotope coded protein labeling (ICPL)1 with immunoprecipitation (IP) and quantitative mass spectrometry (MS). ICPL-IP allows sensitive and accurate analysis of protein interactions from primary tissue. We applied ICPL-IP to immuno-isolate protein complexes from bovine retinal tissue. Protein complexes of immunoprecipitated β-tubulin, a highly abundant protein with known interactors as well as the lowly expressed small GTPase RhoA were analyzed. The results of both analyses demonstrate sensitive and selective identification of known as well as new protein interactions by our method. PMID:23268931

  11. Isotopic exchange between CO2 and H2O and labelling kinetics of photosynthetic oxygen

    International Nuclear Information System (INIS)

    Gerster, Richard

    1971-01-01

    The reaction of carbon dioxide with water has been studied by measuring the rate of oxygen exchange between C 18 O 2 and H 2 16 O. The mathematical treatment of the kinetics allows to determine with accuracy the diffusion flow between the gas and the liquid phase, in the same way as the CO 2 hydration rate. The velocity constant of this last process, whose value gives the in situ enzymatic activity of carbonic anhydrase, has been established in the case of chloroplast and Euglena suspensions and of aerial leaves. The study of the isotopic exchange between C 18 O 2 and a vegetable submitted to alternations of dark and light has allowed to calculate the isotopic abundance of the metabolized CO 2 whose value has been compared to that of the intracellular water and that of photosynthetic oxygen. In addition, a new method using 13 C 18 O 2 gives the means to measure with accuracy eventual isotopic effects. The labelling kinetics of the oxygen evolved by Euglena suspensions whose water has been enriched with 18 O have been established at different temperatures. (author) [fr

  12. Fatty acids labelled in the. omega. -position with iodine isotopes

    Energy Technology Data Exchange (ETDEWEB)

    Mathieu, J.P.; Busquet, G.; Comet, M. (Universite Scientifique et Medicale de Grenoble, 38 - La Tronche (France)); Riche, F.; Vidal, M. (Laboratoire d' Etudes Dynamiques et Structurales de la Selectivite, 38 - Grenoble (France)); Coornaert, S.; Bardy, A. (CEA, Centre de Saclay, 91 - Gif-sur-Yvette (France)); Godart, J. (Institut des Sciences Nucleaires, 38 - Grenoble (France))

    1982-01-01

    The synthesis of saturated acetylenic and olefinic (Z or E) ..omega..-iodinated fatty acids has been carried out and their labelling with iodine-131 or 123 by exchange I/sup -/, *I/sup -/ has been studied. The influence of several parameters -water and fatty acid concentrations, specific activity, labelling solution acidity, iodine carrier presence- on this exchange reaction has been noted, enabling experimental conditions to be defined that produce labelling yields of greater than 95%. These results should lead to widespread clinical use of iodine labelled fatty acids.

  13. Copper absorption from foods labelled intrinsically and extrinsically with Cu-65 stable isotope.

    Science.gov (United States)

    Harvey, L J; Dainty, J R; Beattie, J H; Majsak-Newman, G; Wharf, S G; Reid, M D; Fairweather-Tait, S J

    2005-03-01

    To determine copper absorption from copper containing foods labelled either intrinsically or extrinsically with a highly enriched Cu-65 stable isotope label. A longitudinal cross-over study. The study was conducted at the Institute of Food Research, Human Nutrition Unit, Norwich, UK. Subjects were recruited locally via advertisements placed around the Norwich Research Park. A total of 10 volunteers (nine female, one male) took part in the study, but not all volunteers completed each of the test meals. A highly enriched Cu-65 stable isotope label was administered to volunteers in the form of a reference dose or in breakfast test meals consisting of red wine, soya beans, mushrooms or sunflower seeds. Faecal monitoring and mass spectrometry techniques were used to estimate the relative quantities of copper absorbed from the different test meals. True copper absorption from the reference dose (54%) was similar to extrinsically labelled red wine (49%) and intrinsically labelled sunflower seeds (52%), but significantly higher than extrinsically labelled mushrooms (35%), intrinsically (29%) and extrinsically (15%) labelled soya beans and extrinsically labelled sunflower seed (32%) test meals. The use of Cu-65 extrinsic labels in copper absorption studies requires validation according to the food being examined; intrinsic and extrinsic labelling produced significantly different results for sunflower seeds.

  14. Isotope angiocardiography. Method and preliminary own studies

    Energy Technology Data Exchange (ETDEWEB)

    Stepinska, J; Ruzyllo, W; Konieczny, W [Centrum Medyczne Ksztalcenia Podyplomowego, Warsaw (Poland)

    1979-01-01

    Method of technetium isotope 99 m pass through the heart recording with the aid of radioisotope scanner connected with seriograph and computer is being presented. Preliminary tests were carried out in 26 patients with coronary disease without or with previous myocardial infarction, cardiomyopathy, ventricular septal defect and in patients with artificial mitral and aortic valves. The obtained scans were evaluated qualitatively and compared with performed later contrast X-rays of the heart. Size of the right ventricle, volume and rate of left atrial evacuation, size and contractability of left ventricle were evaluated. Similarity of direct and isotope angiocardiographs, non-invasional character and repeatability of isotope angiocardiography advocate its usefulness.

  15. IsoMS: automated processing of LC-MS data generated by a chemical isotope labeling metabolomics platform.

    Science.gov (United States)

    Zhou, Ruokun; Tseng, Chiao-Li; Huan, Tao; Li, Liang

    2014-05-20

    A chemical isotope labeling or isotope coded derivatization (ICD) metabolomics platform uses a chemical derivatization method to introduce a mass tag to all of the metabolites having a common functional group (e.g., amine), followed by LC-MS analysis of the labeled metabolites. To apply this platform to metabolomics studies involving quantitative analysis of different groups of samples, automated data processing is required. Herein, we report a data processing method based on the use of a mass spectral feature unique to the chemical labeling approach, i.e., any differential-isotope-labeled metabolites are detected as peak pairs with a fixed mass difference in a mass spectrum. A software tool, IsoMS, has been developed to process the raw data generated from one or multiple LC-MS runs by peak picking, peak pairing, peak-pair filtering, and peak-pair intensity ratio calculation. The same peak pairs detected from multiple samples are then aligned to produce a CSV file that contains the metabolite information and peak ratios relative to a control (e.g., a pooled sample). This file can be readily exported for further data and statistical analysis, which is illustrated in an example of comparing the metabolomes of human urine samples collected before and after drinking coffee. To demonstrate that this method is reliable for data processing, five (13)C2-/(12)C2-dansyl labeled metabolite standards were analyzed by LC-MS. IsoMS was able to detect these metabolites correctly. In addition, in the analysis of a (13)C2-/(12)C2-dansyl labeled human urine, IsoMS detected 2044 peak pairs, and manual inspection of these peak pairs found 90 false peak pairs, representing a false positive rate of 4.4%. IsoMS for Windows running R is freely available for noncommercial use from www.mycompoundid.org/IsoMS.

  16. Formation of Hydroxymethyl DNA Adducts in Rats Orally Exposed to Stable Isotope Labeled Methanol

    Science.gov (United States)

    Lu, Kun; Gul, Husamettin; Upton, Patricia B.; Moeller, Benjamin C.; Swenberg, James A.

    2012-01-01

    Methanol is a large volume industrial chemical and widely used solvent and fuel additive. Methanol’s well known toxicity and use in a wide spectrum of applications has raised long-standing environmental issues over its safety, including its carcinogenicity. Methanol has not been listed as a carcinogen by any regulatory agency; however, there are debates about its carcinogenic potential. Formaldehyde, a metabolite of methanol, has been proposed to be responsible for the carcinogenesis of methanol. Formaldehyde is a known carcinogen and actively targets DNA and protein, causing diverse DNA and protein damage. However, formaldehyde-induced DNA adducts arising from the metabolism of methanol have not been reported previously, largely due to the absence of suitable DNA biomarkers and the inability to differentiate what was due to methanol compared with the substantial background of endogenous formaldehyde. Recently, we developed a unique approach combining highly sensitive liquid chromatography-mass spectrometry methods and exposure to stable isotope labeled chemicals to simultaneously quantify formaldehyde-specific endogenous and exogenous DNA adducts. In this study, rats were exposed daily to 500 or 2000 mg/kg [13CD4]-methanol by gavage for 5 days. Our data demonstrate that labeled formaldehyde arising from [13CD4]-methanol induced hydroxymethyl DNA adducts in multiple tissues in a dose-dependent manner. The results also demonstrated that the number of exogenous DNA adducts was lower than the number of endogenous hydroxymethyl DNA adducts in all tissues of rats administered 500 mg/kg per day for 5 days, a lethal dose to humans, even after incorporating an average factor of 4 for reduced metabolism due to isotope effects of deuterium-labeled methanol into account. PMID:22157354

  17. Synthesis of 13C and 2H labelled retinals: spectroscopic investigations on isotopically labelled rhodopsin and bacteriorhodopsin

    International Nuclear Information System (INIS)

    Pardoen, J.A.

    1986-01-01

    In order to develop probes of the structure of chromophores, the author introduces isotopic modifications at specific chromophoric positions as structural probes. To obtain bacteriorhodopsin, rhodopsin and their photoproducts labelled in the chromophore at selected positions, bacterioopsin and opsin were reacted with the appropriate labelled a11-trans and 11-cis retinals. The author describes the synthesis of a11-trans retinal selectively 13 C labelled at different positions. The characterization of these labelled a11-trans retinals by mass spectrometry, 300 MHz 1 H NMR and 75 MHz 13 C NMR spectroscopy is given. The photochemical preparation and isolation of the pure 9-, 11- and 13-cis forms is described in the experimental part. (Auth.)

  18. The labelling of Nanocoll[reg] with [111In] for dual-isotope scanning

    International Nuclear Information System (INIS)

    Mitterhauser, Markus; Wadsak, Wolfgang; Key Mien, Leonhard-; Eidherr, Harald; Roka, Sebastian; Zettinig, Georg; Angelberger, Peter; Viernstein, Helmut; Kletter, Kurt; Dudczak, Robert

    2003-01-01

    Visualization and biopsy of sentinel lymph nodes play an important role in planning and controlling the therapy of breast cancer. Hitherto two methods--scintigraphy or gamma probe detection after injection of [ 99m Tc]-nanocolloids and visual detection after injection of patent blue dye--are used routinely. There are no conclusive publications elucidating such important parameters as injection site, injection method and colloidal parameters. The present work aims to label Nanocoll[reg] with [ 111 In] to provide an alternative method, a simultaneous one-compound dual-isotope application. Methods: [ 111 In]-Indiumchloride was buffered with acetate and transferred to the nanocolloid. The colloid labelling reaction was complete after 30 min and filtrated through 100 nm Nuclepore[reg] filters. Results: Incorporation yield of [ 111 In]-Indium into the nanocolloid was nearly quantitative, the step associated with the major loss of activity was the particle sizing with a mean yield of 55%. Conclusion: The presented method allows for the routine supply of [ 111 In]-nanocolloids. Size-filtered [ 111 In]-Nanocoll[reg] shows the same particle size range as [ 99m Tc]-Nanocoll[reg

  19. Chemical Ligation of Folded Recombinant Proteins: Segmental Isotopic Labeling of Domains for NMR Studies

    Science.gov (United States)

    Xu, Rong; Ayers, Brenda; Cowburn, David; Muir, Tom W.

    1999-01-01

    A convenient in vitro chemical ligation strategy has been developed that allows folded recombinant proteins to be joined together. This strategy permits segmental, selective isotopic labeling of the product. The src homology type 3 and 2 domains (SH3 and SH2) of Abelson protein tyrosine kinase, which constitute the regulatory apparatus of the protein, were individually prepared in reactive forms that can be ligated together under normal protein-folding conditions to form a normal peptide bond at the ligation junction. This strategy was used to prepare NMR sample quantities of the Abelson protein tyrosine kinase-SH(32) domain pair, in which only one of the domains was labeled with 15N Mass spectrometry and NMR analyses were used to confirm the structure of the ligated protein, which was also shown to have appropriate ligand-binding properties. The ability to prepare recombinant proteins with selectively labeled segments having a single-site mutation, by using a combination of expression of fusion proteins and chemical ligation in vitro, will increase the size limits for protein structural determination in solution with NMR methods. In vitro chemical ligation of expressed protein domains will also provide a combinatorial approach to the synthesis of linked protein domains.

  20. Accurate and sensitive determination of molar fractions of "1"3C-Labeled intracellular metabolites in cell cultures grown in the presence of isotopically-labeled glucose

    International Nuclear Information System (INIS)

    Fernández-Fernández, Mario; Rodríguez-González, Pablo; Hevia Sánchez, David; González-Menéndez, Pedro; Sainz Menéndez, Rosa M.; García Alonso, J. Ignacio

    2017-01-01

    This work describes a methodology based on multiple linear regression and GC-MS for the determination of molar fractions of isotopically-labeled intracellular metabolites in cell cultures. Novel aspects of this work are: i) the calculation of theoretical isotopic distributions of the different isotopologues from an experimentally measured value of % 13C enrichment of the labeled precursor ii) the calculation of the contribution of lack of mass resolution of the mass spectrometer and different fragmentation mechanism such as the loss or gain of hydrogen atoms in the EI source to measure the purity of the selected cluster for each metabolite and iii) the validation of the methodology not only by the analysis of gravimetrically prepared mixtures of isotopologues but also by the comparison of the obtained molar fractions with experimental values obtained by GC-Combustion-IRMS based on "1"3C/"1"2C isotope ratio measurements. The method is able to measure molar fractions for twenty-eight intracellular metabolites derived from glucose metabolism in cell cultures grown in the presence of "1"3C-labeled Glucose. The validation strategies demonstrate a satisfactory accuracy and precision of the proposed procedure. Also, our results show that the minimum value of "1"3C incorporation that can be accurately quantified is significantly influenced by the calculation of the spectral purity of the measured cluster and the number of "1"3C atoms of the labeled precursor. The proposed procedure was able to accurately quantify gravimetrically prepared mixtures of natural and labeled glucose molar fractions of 0.07% and mixtures of natural and labeled glycine at molar fractions down to 0.7%. The method was applied to initial studies of glucose metabolism of different prostate cancer cell lines. - Highlights: • Determination of molar fractions of "1"3C-labeled metabolites in cell cultures. • The method is based on multiple linear regression and GC-MS. • Validation of the method by

  1. Accurate and sensitive determination of molar fractions of {sup 13}C-Labeled intracellular metabolites in cell cultures grown in the presence of isotopically-labeled glucose

    Energy Technology Data Exchange (ETDEWEB)

    Fernández-Fernández, Mario [Department of Physical and Analytical Chemistry, Faculty of Chemistry, University of Oviedo, Julián Clavería 8, 33006 Oviedo (Spain); Rodríguez-González, Pablo, E-mail: rodriguezpablo@uniovi.es [Department of Physical and Analytical Chemistry, Faculty of Chemistry, University of Oviedo, Julián Clavería 8, 33006 Oviedo (Spain); Hevia Sánchez, David; González-Menéndez, Pedro; Sainz Menéndez, Rosa M. [University Institute of Oncology (IUOPA), University of Oviedo, Julián Clavería 6, 33006 Oviedo (Spain); García Alonso, J. Ignacio [Department of Physical and Analytical Chemistry, Faculty of Chemistry, University of Oviedo, Julián Clavería 8, 33006 Oviedo (Spain)

    2017-05-29

    This work describes a methodology based on multiple linear regression and GC-MS for the determination of molar fractions of isotopically-labeled intracellular metabolites in cell cultures. Novel aspects of this work are: i) the calculation of theoretical isotopic distributions of the different isotopologues from an experimentally measured value of % 13C enrichment of the labeled precursor ii) the calculation of the contribution of lack of mass resolution of the mass spectrometer and different fragmentation mechanism such as the loss or gain of hydrogen atoms in the EI source to measure the purity of the selected cluster for each metabolite and iii) the validation of the methodology not only by the analysis of gravimetrically prepared mixtures of isotopologues but also by the comparison of the obtained molar fractions with experimental values obtained by GC-Combustion-IRMS based on {sup 13}C/{sup 12}C isotope ratio measurements. The method is able to measure molar fractions for twenty-eight intracellular metabolites derived from glucose metabolism in cell cultures grown in the presence of {sup 13}C-labeled Glucose. The validation strategies demonstrate a satisfactory accuracy and precision of the proposed procedure. Also, our results show that the minimum value of {sup 13}C incorporation that can be accurately quantified is significantly influenced by the calculation of the spectral purity of the measured cluster and the number of {sup 13}C atoms of the labeled precursor. The proposed procedure was able to accurately quantify gravimetrically prepared mixtures of natural and labeled glucose molar fractions of 0.07% and mixtures of natural and labeled glycine at molar fractions down to 0.7%. The method was applied to initial studies of glucose metabolism of different prostate cancer cell lines. - Highlights: • Determination of molar fractions of {sup 13}C-labeled metabolites in cell cultures. • The method is based on multiple linear regression and GC-MS.

  2. Isotopic methods or immuno diagnosis: The Radioimmunoassay and immunoradiometric assay

    International Nuclear Information System (INIS)

    Caso, R.

    1997-01-01

    This work offers an explanation about the more used isotopic techniques for immuno diagnosis: the radioimmunoassay (RIA) and immunoradiometric assay (IRMA). It describes the basic principles of these assays, the antigen-antibody reaction, the radioiodination methods with I-125 for antigens and antibodies, the purification and characterization of labelled compounds. On the order hand they present work gives a review of the methods for separate the bound and free fractions. At the end it offers the principles of the quality control of immunoassay and the future lines of research in the field of RIA and IRMA

  3. Existing and emerging technologies for measuring stable isotope labelled retinol in biological samples: isotope dilution analysis of body retinol stores.

    Science.gov (United States)

    Preston, Tom

    2014-01-01

    This paper discusses some of the recent improvements in instrumentation used for stable isotope tracer measurements in the context of measuring retinol stores, in vivo. Tracer costs, together with concerns that larger tracer doses may perturb the parameter under study, demand that ever more sensitive mass spectrometric techniques are developed. GCMS is the most widely used technique. It has high sensitivity in terms of sample amount and uses high resolution GC, yet its ability to detect low isotope ratios is limited by background noise. LCMSMS may become more accessible for tracer studies. Its ability to measure low level stable isotope tracers may prove superior to GCMS, but it is isotope ratio MS (IRMS) that has been designed specifically for low level stable isotope analysis through accurate analysis of tracer:tracee ratios (the tracee being the unlabelled species). Compound-specific isotope analysis, where GC is interfaced to IRMS, is gaining popularity. Here, individual 13C-labelled compounds are separated by GC, combusted to CO2 and transferred on-line for ratiometric analysis by IRMS at the ppm level. However, commercially-available 13C-labelled retinol tracers are 2 - 4 times more expensive than deuterated tracers. For 2H-labelled compounds, GC-pyrolysis-IRMS has now become more generally available as an operating mode on the same IRMS instrument. Here, individual compounds are separated by GC and pyrolysed to H2 at high temperature for analysis by IRMS. It is predicted that GC-pyrolysis-IRMS will facilitate low level tracer procedures to measure body retinol stores, as has been accomplished in the case of fatty acids and amino acids. Sample size requirements for GC-P-IRMS may exceed those of GCMS, but this paper discusses sample preparation procedures and predicts improvements, particularly in the efficiency of sample introduction.

  4. The method of obtaining of sodium orthoiodohippurate labelled with iodine-131

    International Nuclear Information System (INIS)

    Aripov, D.; Abdukayumov, M.; Shukurov, A.Sh.

    1994-01-01

    The method of labelling of sodium orthoiodohippurate was elaborated with the purpose of increasing the preparation quality. Method includes the reaction of isotopic exchange between orthoiodhippur acid and sodium iodide solution labelled with iodine-131 with volume activity 150-200 mCu/mL and pH=6,5-7,0. Reaction occurs at temperature 120-130 C during 1,1-1,3 hours and the compound obtained is dissolved in 1% sodium bicarbonate solution. (author)

  5. Kinetic isotope effects in reaction of ferment oxidation of tritium-labelled D-galactosamine

    International Nuclear Information System (INIS)

    Akulov, G.P.; Korsakova, N.A.

    1992-01-01

    Primary, secondary and intramolecular kinetic isotopic effects in reaction of ferment oxidation of D-galactosamine labelled by tritium in position 6, were measured. When comparing values of the effects with previously obtained results for similar reaction D-[6- 3 H]galactose, it was ascertained that the presence of aminogroup in galactopyranosyl mainly affects kinetics of substrate-ferment complex formation stage. The possibility to use kinetic isotope effects for increase in molar activity of D-galactosamine, labelled by tritium in position 6, is shown

  6. Negative ion ESI-MS analysis of natural yellow dye flavonoids--An isotopic labelling study

    Science.gov (United States)

    McNab, Hamish; Ferreira, Ester S. B.; Hulme, Alison N.; Quye, Anita

    2009-07-01

    Flavonoids are amongst the most commonly used natural yellow colourants in paintings, as lakes, and in historical textiles as mordant dyes. In this paper, evidence from isotopically labelled substrates is used to propose negative ion electrospray collision induced decomposition mechanisms of flavones, flavonols and an isoflavone. These mechanisms include a retro-Diels-Alder fragmentation (observed for flavones and flavonols) and an M-122 fragmentation (characteristic of 3',4'-dihydroxyflavonols). In addition, the presence of a m/z 125 fragment ion is shown to be characteristic of 2'-hydroxyflavonols and an ion at m/z 149 is shown to be characteristic of 4'-hydroxyflavones. Applications of these methods are exemplified by the identification of a minor component of Dyer's camomile (Anthemis tinctoria L.) and the identification of the dye source in green threads sampled from an 18th Century Scottish tartan fragment.

  7. Direct isotope determination of isotopically labelled lipids by field desorption mass spectrometry

    International Nuclear Information System (INIS)

    Lehmann, W.D.; Kessler, M.

    1982-01-01

    Lipids labelled with deuterium or carbon-14 have been investigated by field desorption mass spectrometry for determination of their degree of labelling. This application is demonstrated for free fatty acids, cholesterol, cholesteryl esters, triglycerides, and L-α-phosphatidylcholines. Comparison of the molecular ion groups of the non-labelled and of the labelled compounds enables a fast and reliable determination of the degree of labelling. For multiply labelled compounds the label distribution is also obtained from the molecular ion group. In addition, for cholesteryl esters and for phosphatidylcholines structurally significant fragment ions provide information about the position of the label. Several hundred nanograms of the compound are typically required for a single analysis with a relative standard error of 0.5-2% in the value calculated for atom% hydrogen-2 or for the specific carbon-14 activity. (orig.) [de

  8. Affordable uniform isotope labeling with 2H, 13C and 15N in insect cells

    International Nuclear Information System (INIS)

    Sitarska, Agnieszka; Skora, Lukasz; Klopp, Julia; Roest, Susan; Fernández, César; Shrestha, Binesh; Gossert, Alvar D.

    2015-01-01

    For a wide range of proteins of high interest, the major obstacle for NMR studies is the lack of an affordable eukaryotic expression system for isotope labeling. Here, a simple and affordable protocol is presented to produce uniform labeled proteins in the most prevalent eukaryotic expression system for structural biology, namely Spodoptera frugiperda insect cells. Incorporation levels of 80 % can be achieved for 15 N and 13 C with yields comparable to expression in full media. For 2 H, 15 N and 2 H, 13 C, 15 N labeling, incorporation is only slightly lower with 75 and 73 %, respectively, and yields are typically twofold reduced. The media were optimized for isotope incorporation, reproducibility, simplicity and cost. High isotope incorporation levels for all labeling patterns are achieved by using labeled algal amino acid extracts and exploiting well-known biochemical pathways. The final formulation consists of just five commercially available components, at costs 12-fold lower than labeling media from vendors. The approach was applied to several cytosolic and secreted target proteins

  9. A free-air system for long-term stable carbon isotope labeling of adult forest trees

    Science.gov (United States)

    Stable carbon (C) isotopes, in particular employed in labeling experiments, are an ideal tool to broaden our understanding of C dynamics in trees and forest ecosystems. Here, we present a free-air exposure system, named isoFACE, designed for long-term stable C isotope labeling in...

  10. Some methods for labelling organic compounds by deuterium

    International Nuclear Information System (INIS)

    Moustapha, C.

    1988-01-01

    The rapid growth of knowledge in the fields of biochemistry, physiology, and molecular biology reflects to a considerable degree the utilization of stable isotopes (specially deuterium) in the study of chemical reactions and fragmentation mechanisms in mass spectrometry, as well as in the pharmacological and biological studies. Organic compounds maybe labelled by deuterium through classic organic reactions by using special deuterated solvents and reagents. This article discusses some reactions, with examples on how to prepare labelled compounds with high isotopic purety. These reactions are: exchange reactions in acid and alkaline media (the exchange in the chromatographic column in liquid and gas phases, the exchange in homogenous medium), reduction reactions of functional groups as well as saturation of the double bounds by deuterium using hydrogenation catalystes, electrochemical reactions using KOLBE, and photochemical reactions. This article also deals with spectroscopic properties of deuterium and the methods which are used to identify its compounds such as infrared, nuclear magnetic resonance, and mass spectroscopy. 37 refs., 2 figs

  11. Method of eliminating gaseous hydrogen isotopes

    International Nuclear Information System (INIS)

    Nagakura, Masaaki; Imaizumi, Hideki; Suemori, Nobuo; Aizawa, Takashi; Naito, Taisei.

    1983-01-01

    Purpose: To prevent external diffusion of gaseous hydrogen isotopes such as tritium or the like upon occurrence of tritium leakage accident in a thermonuclear reactor by recovering to eliminate the isotopes rapidly and with safety. Method: Gases at the region of a reactor container where hydrogen isotopes might leak are sucked by a recycing pump, dehumidified in a dehumidifier and then recycled from a preheater through a catalytic oxidation reactor to a water absorption tower. In this structure, the dehumidifier is disposed at the upstream of the catalytic oxidation reactor to reduce the water content of the gases to be processed, whereby the eliminating efficiency for the gases to be processed can be maintained well even when the oxidation reactor is operated at a low temperature condition near the ambient temperature. This method is based on the fact that the oxidating reactivity of the catalyst can be improved significantly by eliminating the water content in the gases to be processed. (Yoshino, Y.)

  12. Hydrogen isotope separation by cryogenic distillation method

    International Nuclear Information System (INIS)

    Hayakawa, Nobuo; Mitsui, Jin

    1987-01-01

    Hydrogen isotope separation in fusion fuel cycle and tritium recovery from heavy water reactor are very important, and therefore the early establishment of these separation techniques are desired. The cryogenic distillation method in particular is promising for the separation of hydrogen isotope and the recovery of high concentrated tritium. The studies of hydrogen isotope separation by cryogenic distillation method have been carried out by using the experimental apparatus made for the first time in Japan. The separation of three components (H 2 -HD-D 2 ) under total reflux conditions was got by using the packing tower of 500 mm height. It was confirmed that the Height Equivalent Theoretical Plate (HETP) was 20 - 30 mm for the vapor's line velocity of 20 - 80 mm/s. (author)

  13. Synthesis of stable isotopically labeled peptides with filter-assisted enzymatic labeling for the diagnosis of hepatitis B virus infection utilizing mass spectrometry-based proteomics strategy

    International Nuclear Information System (INIS)

    Tsai, Hsing-Fen; Hsiao, He-Hsuan

    2017-01-01

    A facile method for the preparation of stable isotopically labeled peptides was developed by means of filter-assisted tryptic "1"6O/"1"8O water labeling, which could be directly applied to the determination of hepatitis B virus infection from human serum with tandem mass spectrometry. Tryptic peptides of hepatitis B surface antigen or hepatitis B e antigen from different subtypes of hepatitis B virus were synthesized with traditional solid-phase peptide synthesis as potential biomarkers. Trypsin catalyzed oxygen-18 exchange at their amidated c-terminus of arginine or lysine residue. The protease catalyzed oxygen-18 to oxygen-16 back exchange reaction was eliminated due to the complete removal of trypsin by the centrifugal filter containing a thin membrane associated with molecular weight cut-off of 10 KDa. The synthetic isotopic peptides were spiked into trichloroacetic acid/acetone precipitated human serum as internal standards and were selectively detected with multiplexed parallel reaction monitoring on a hybrid quadrupole-orbitrap mass spectrometer. The limit of detection for all synthetic peptides were in the range of 0.09 fmol–1.13 fmol. The results indicated that the peptide YLWEWASVR derived from hepatitis B surface antigen was quantified approximately 200 fmol per μl serum and may serve as a diagnostic biomarker for the detection of hepatitis B virus infected disease. - Highlights: • Facile synthesis of an inexpensive and highly reproducible stable isotopically labeled peptides. • Complete incorporation of two "1"8O atoms into synthesized peptides with filter-assisted enzymatic labeling. • Targeted analysis with parallel reaction monitoring assay for the disease diagnosis.

  14. Synthesis of stable isotopically labeled peptides with filter-assisted enzymatic labeling for the diagnosis of hepatitis B virus infection utilizing mass spectrometry-based proteomics strategy

    Energy Technology Data Exchange (ETDEWEB)

    Tsai, Hsing-Fen; Hsiao, He-Hsuan, E-mail: hhhsiao@dragon.nchu.edu.tw

    2017-03-01

    A facile method for the preparation of stable isotopically labeled peptides was developed by means of filter-assisted tryptic {sup 16}O/{sup 18}O water labeling, which could be directly applied to the determination of hepatitis B virus infection from human serum with tandem mass spectrometry. Tryptic peptides of hepatitis B surface antigen or hepatitis B e antigen from different subtypes of hepatitis B virus were synthesized with traditional solid-phase peptide synthesis as potential biomarkers. Trypsin catalyzed oxygen-18 exchange at their amidated c-terminus of arginine or lysine residue. The protease catalyzed oxygen-18 to oxygen-16 back exchange reaction was eliminated due to the complete removal of trypsin by the centrifugal filter containing a thin membrane associated with molecular weight cut-off of 10 KDa. The synthetic isotopic peptides were spiked into trichloroacetic acid/acetone precipitated human serum as internal standards and were selectively detected with multiplexed parallel reaction monitoring on a hybrid quadrupole-orbitrap mass spectrometer. The limit of detection for all synthetic peptides were in the range of 0.09 fmol–1.13 fmol. The results indicated that the peptide YLWEWASVR derived from hepatitis B surface antigen was quantified approximately 200 fmol per μl serum and may serve as a diagnostic biomarker for the detection of hepatitis B virus infected disease. - Highlights: • Facile synthesis of an inexpensive and highly reproducible stable isotopically labeled peptides. • Complete incorporation of two {sup 18}O atoms into synthesized peptides with filter-assisted enzymatic labeling. • Targeted analysis with parallel reaction monitoring assay for the disease diagnosis.

  15. Stable isotope labeling – Liquid chromatography/mass spectrometry for quantitative analysis of androgenic and progestagenic steroids

    International Nuclear Information System (INIS)

    Guo, Ning; Liu, Ping; Ding, Jun; Zheng, Shu-Jian; Yuan, Bi-Feng; Feng, Yu-Qi

    2016-01-01

    Steroid hormones play important roles in mammal at very low concentrations and are associated with numerous endocrinology and oncology diseases. Therefore, quantitative analysis of steroid hormones can provide crucial information for uncovering underlying mechanisms of steroid hormones related diseases. In the current study, we developed a sensitive method for the detection of steroid hormones (progesterone, dehydroepiandrosterone, testosterone, pregnenolone, 17-hydroxyprogesterone, androstenedione and 17α-hydroxypregnenolone) in body fluids by stable isotope labeling coupled with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) analysis. In this respect, a pair of isotopes labeling reagents, Girard reagent P (GP) and d_5-Girard reagent P (d_5-GP), were synthesized and utilized to label steroid hormones in follicular fluid samples and steroid hormone standards, respectively. The heavy labeled standards were used as internal standards for quantification to minimize quantitation deviation in MS analysis due to the matrix and ion suppression effects. The ionization efficiencies of steroid hormones were greatly improved by 4–504 folds through the introduction of a permanent charged moiety of quaternary ammonium from GP. Using the developed method, we successfully quantified steroid hormones in human follicular fluid. We found that the contents of testosterone and androstenedione exhibited significant increase while the content of pregnenolone had significant decrease in follicular fluid of polycystic ovarian syndrome (PCOS) patients compared with healthy controls, indicating that these steroid hormones with significant change may contribute to the pathogenesis of PCOS. Taken together, the developed stable isotope labeling coupled LC-ESI-MS/MS analysis demonstrated to be a promising method for the sensitive and accurate determination of steroid hormones, which may facilitate the in-depth investigation of steroid hormones related

  16. Stable isotope labeling – Liquid chromatography/mass spectrometry for quantitative analysis of androgenic and progestagenic steroids

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Ning; Liu, Ping; Ding, Jun; Zheng, Shu-Jian; Yuan, Bi-Feng; Feng, Yu-Qi, E-mail: yqfeng@whu.edu.cn

    2016-01-28

    Steroid hormones play important roles in mammal at very low concentrations and are associated with numerous endocrinology and oncology diseases. Therefore, quantitative analysis of steroid hormones can provide crucial information for uncovering underlying mechanisms of steroid hormones related diseases. In the current study, we developed a sensitive method for the detection of steroid hormones (progesterone, dehydroepiandrosterone, testosterone, pregnenolone, 17-hydroxyprogesterone, androstenedione and 17α-hydroxypregnenolone) in body fluids by stable isotope labeling coupled with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) analysis. In this respect, a pair of isotopes labeling reagents, Girard reagent P (GP) and d{sub 5}-Girard reagent P (d{sub 5}-GP), were synthesized and utilized to label steroid hormones in follicular fluid samples and steroid hormone standards, respectively. The heavy labeled standards were used as internal standards for quantification to minimize quantitation deviation in MS analysis due to the matrix and ion suppression effects. The ionization efficiencies of steroid hormones were greatly improved by 4–504 folds through the introduction of a permanent charged moiety of quaternary ammonium from GP. Using the developed method, we successfully quantified steroid hormones in human follicular fluid. We found that the contents of testosterone and androstenedione exhibited significant increase while the content of pregnenolone had significant decrease in follicular fluid of polycystic ovarian syndrome (PCOS) patients compared with healthy controls, indicating that these steroid hormones with significant change may contribute to the pathogenesis of PCOS. Taken together, the developed stable isotope labeling coupled LC-ESI-MS/MS analysis demonstrated to be a promising method for the sensitive and accurate determination of steroid hormones, which may facilitate the in-depth investigation of steroid hormones

  17. Expeditious syntheses of stable and radioactive isotope-labeled anticonvulsant agent, JNJ-26990990, and its metabolites.

    Science.gov (United States)

    Lin, Ronghui; Weaner, Larry E; Hoerr, David C; Salter, Rhys; Gong, Yong

    2013-01-01

    Syntheses of stable and radioactive isotope-labeled anticonvulsant agent, JNJ-26990990, that is, N-(benzo[b]thien-3-ylmethyl)-sulfamide and its metabolites are described. [(13)C(15)N]Benzo[b]thiophene-3-carbonitrile was first prepared by coupling of 3-bromo-benzo[b]thiophene with [(13)C(15)N]-copper cyanide. The resultant [(13)C(15)N]benzo[b]thiophene-3-carbonitrile was reduced with lithium aluminum deuteride to give [(13)CD2(15)N]benzo[b]thiophen-3-yl-methylamine; which was then coupled with sulfamide to afford [(13)CD2(15)N]-N-(benzo[b]thien-3-ylmethyl)-sulfamide, the stable isotope-labeled compound with four stable isotope atoms. Direct oxidation of [(13)CD2(15)N]-N-(benzo[b]thien-3-ylmethyl)-sulfamide with hydrogen peroxide and peracetic acid gave the stable isotope-labeled sulfoxide and sulfone metabolites. On the other hand, radioactive (14)C-labeled N-(benzo[b]thien-3-ylmethyl)-sulfamide was prepared conveniently by sequential coupling of 3-bromo-benzo[b]thiophene with [(14)C]-copper cyanide, reduction of the carbonitrile to carboxaldehyde, and reductive amination with sulfamide. Copyright © 2013 John Wiley & Sons, Ltd.

  18. Addressing Raman features of individual layers in isotopically labeled Bernal stacked bilayer graphene

    Czech Academy of Sciences Publication Activity Database

    da Costa, Sara; Ek Weis, Johan; Frank, Otakar; Fridrichová, Michaela; Kalbáč, Martin

    2016-01-01

    Roč. 3, č. 2 (2016), 025022 ISSN 2053-1583 R&D Projects: GA MŠk LL1301 Institutional support: RVO:61388955 Keywords : graphene bilayer * Raman spectroscopy * isotope labeling Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 6.937, year: 2016

  19. Identification of miRNA targets with stable isotope labeling by amino acids in cell culture

    DEFF Research Database (Denmark)

    Vinther, Jeppe; Hedegaard, Mads Marquardt; Gardner, Paul Phillip

    2006-01-01

    miRNAs are small noncoding RNAs that regulate gene expression. We have used stable isotope labeling by amino acids in cell culture (SILAC) to investigate the effect of miRNA-1 on the HeLa cell proteome. Expression of 12 out of 504 investigated proteins was repressed by miRNA-1 transfection...

  20. Temperature-induced strain and doping in monolayer and bilayer isotopically labeled graphene

    Czech Academy of Sciences Publication Activity Database

    Verhagen, Timotheus; Drogowska, Karolina; Kalbáč, Martin; Vejpravová, Jana

    2015-01-01

    Roč. 92, č. 12 (2015), "125437-1"-"125437-9" ISSN 1098-0121 R&D Projects: GA ČR GA15-02196S; GA MŠk LL1301 Institutional support: RVO:68378271 ; RVO:61388955 Keywords : isotopically labeled graphene * temperature dependence * Raman spectroscopy * phonons Subject RIV: BM - Solid Matter Physics ; Magnetism Impact factor: 3.736, year: 2014

  1. Isotope effect study of κ-(BEDT-TTF)2Cu(NCS)2: Labeling in the anion

    International Nuclear Information System (INIS)

    Kini, A.M.; Wang, H.H.; Schlueter, J.A.

    1995-01-01

    Since the initial discovery of organic superconductivity in 1979, a large number of organic superconductors have now been synthesized. However, the mechanism of electron-pairing in these novel superconductors has remained largely unresolved. Isotope effect studies constitute an important experimental tool for the investigation of whether or not the electron-pairing mechanism in organic superconductors is phonon-mediated, as in conventional superconductors. Recent isotope effect studies in the authors' laboratory, involving seven different isotopically labeled BEDT-TTF (or ET) derivatives, have demonstrated the following: (1) intramolecular phonon modes involving C double-bond C and Csingle bondS stretching vibrations in the ET donor molecule are not the dominant mediators of electron-pairing, and (2) in κ-(ET) 2 Cu(NCS) 2 , there exist two competing isotope effects--a normal mass effect, i.e., lowering of T c upon isotopic labeling, when the ET molecular mass is increased by concurrent 13 C and 34 S labeling, in addition to an inverse isotope effect upon deuterium labeling in ET. It is of great interest to investigate if there is an isotope effect when the charge-compensating anions, which are also located within the non-conducting layer in the superconducting cation-radical salts, are isotopically labeled. The existence of an isotope effect when the anions are labeled would be indicative of electron-pairing with the mediation of vibrational frequencies associated with the anions. In this paper, the authors present the results of the first isotope effect study in which isotopic labeling in the anion portion of κ-(ET) 2 Cu(NCS) 2 is carried out. The authors find no isotope effect when the carbon and nitrogen atoms of the thiocyanate groups in the anion are replaced with 13 C and 15 N isotopes

  2. Cell-free expression and stable isotope labelling strategies for membrane proteins

    International Nuclear Information System (INIS)

    Sobhanifar, Solmaz; Reckel, Sina; Junge, Friederike; Schwarz, Daniel; Kai, Lei; Karbyshev, Mikhail; Loehr, Frank; Bernhard, Frank; Doetsch, Volker

    2010-01-01

    Membrane proteins are highly underrepresented in the structural data-base and remain one of the most challenging targets for functional and structural elucidation. Their roles in transport and cellular communication, furthermore, often make over-expression toxic to their host, and their hydrophobicity and structural complexity make isolation and reconstitution a complicated task, especially in cases where proteins are targeted to inclusion bodies. The development of cell-free expression systems provides a very interesting alternative to cell-based systems, since it circumvents many problems such as toxicity or necessity for the transportation of the synthesized protein to the membrane, and constitutes the only system that allows for direct production of membrane proteins in membrane-mimetic environments which may be suitable for liquid state NMR measurements. The unique advantages of the cell-free expression system, including strong expression yields as well as the direct incorporation of almost any combination of amino acids with very little metabolic scrambling, has allowed for the development of a wide-array of isotope labelling techniques which facilitate structural investigations of proteins whose spectral congestion and broad line-widths may have earlier rendered them beyond the scope of NMR. Here we explore various labelling strategies in conjunction with cell-free developments, with a particular focus on α-helical transmembrane proteins which benefit most from such methods.

  3. Differential Isotope Labeling of Glycopeptides for Accurate Determination of Differences in Site-Specific Glycosylation.

    Science.gov (United States)

    Pabst, Martin; Benešová, Iva; Fagerer, Stephan R; Jacobsen, Mathias; Eyer, Klaus; Schmidt, Gregor; Steinhoff, Robert; Krismer, Jasmin; Wahl, Fabian; Preisler, Jan; Zenobi, Renato

    2016-01-04

    We introduce a stable isotope labeling approach for glycopeptides that allows a specific glycosylation site in a protein to be quantitatively evaluated using mass spectrometry. Succinic anhydride is used to specifically label primary amino groups of the peptide portion of the glycopeptides. The heavy form (D4(13)C4) provides an 8 Da mass increment over the light natural form (H4(12)C4), allowing simultaneous analysis and direct comparison of two glycopeptide profiles in a single MS scan. We have optimized a protocol for an in-solution trypsin digestion, a one-pot labeling procedure, and a post-labeling solid-phase extraction to obtain purified and labeled glycopeptides. We provide the first demonstration of this approach by comparing IgG1 Fc glycopeptides from polyclonal IgG samples with respect to their galactosylation and sialylation patterns using MALDI MS and LC-ESI-MS.

  4. Assessing wine quality using isotopic methods

    International Nuclear Information System (INIS)

    Costinel, Diana; Ionete, Roxana Elena; Vremera, Raluca; Stefanescu, Ioan

    2010-01-01

    Full text: The analytical methods used to determine the isotope ratios of deuterium, carbon-13 and oxygen-18 in wines have gained official recognition from the Office International de la Vigne et du Vin (OIV) and National Organisation of Vine and Wine. The amount of stable isotopes in water and carbon dioxide from plant organic materials and their distribution in sugar and ethanol molecules are influenced by geo-climatic conditions of the region, grape varieties and the year of harvest. For wine characterization, to prove the botanical and geographical origin of the raw material, the isotopic analysis by continuous flow mass spectrometry CF-IRMS has made a significant contribution. This paper emphasize the results of a study concerning the assessing of water adulterated wines and non-grape alcohol and sugar additions at different concentration levels, using CF-IRMS analytical technique. (authors)

  5. Stable-isotope-labeled carbohydrates and nucleosides: Synthesis and applications in chemistry and biology

    International Nuclear Information System (INIS)

    Serianni, A.S.

    1994-01-01

    Carbohydrates play important roles in many key biochemical processes in living cells. For example, they are metabolized to produce energy, mediate cell-cell recognition, and play an indirect role (as constituents of DNA and RNA) in DNA replication, RNA transcription, and protein synthesis. These roles, and others of comparable biochemical significance, have been studied to varying extends with the use of stable isotopically labeled molecules, usually in conjunction with NMR spectroscopy and/or mass spectrometry. For example, carbohydrate metabolism has been monitored in vitro and in vivo with the use of isotopically labeled compounds. Molecular aspects of cell-cell recognition, mediated by cell-surface glycoproteins and glycolipids, have been probed through NMR studies of isotopically labeled oligosaccharides. More recently, the solution behavior of DNA and RNA has been examined through the use of labeled oligonucleotides. In all of these pursuits, the effort and expense to prepare labeled molecules, both of which can be substantial, are more than offset by the wealth of information derived from these studies. This information often cannot be accessed, or can be accessed only with great difficulty, using natural (unlabeled) compounds

  6. Stable-isotope-labeled carbohydrates and nucleosides: Synthesis and applications in chemistry and biology

    Energy Technology Data Exchange (ETDEWEB)

    Serianni, A.S. [Univ. of Notre Dame, IN (United States)

    1994-12-01

    Carbohydrates play important roles in many key biochemical processes in living cells. For example, they are metabolized to produce energy, mediate cell-cell recognition, and play an indirect role (as constituents of DNA and RNA) in DNA replication, RNA transcription, and protein synthesis. These roles, and others of comparable biochemical significance, have been studied to varying extends with the use of stable isotopically labeled molecules, usually in conjunction with NMR spectroscopy and/or mass spectrometry. For example, carbohydrate metabolism has been monitored in vitro and in vivo with the use of isotopically labeled compounds. Molecular aspects of cell-cell recognition, mediated by cell-surface glycoproteins and glycolipids, have been probed through NMR studies of isotopically labeled oligosaccharides. More recently, the solution behavior of DNA and RNA has been examined through the use of labeled oligonucleotides. In all of these pursuits, the effort and expense to prepare labeled molecules, both of which can be substantial, are more than offset by the wealth of information derived from these studies. This information often cannot be accessed, or can be accessed only with great difficulty, using natural (unlabeled) compounds.

  7. Hydroponic isotope labeling of entire plants and high-performance mass spectrometry for quantitative plant proteomics.

    Science.gov (United States)

    Bindschedler, Laurence V; Mills, Davinia J S; Cramer, Rainer

    2012-01-01

    Hydroponic isotope labeling of entire plants (HILEP) combines hydroponic plant cultivation and metabolic labeling with stable isotopes using (15)N-containing inorganic salts to label whole and mature plants. Employing (15)N salts as the sole nitrogen source for HILEP leads to the production of healthy-looking plants which contain (15)N proteins labeled to nearly 100%. Therefore, HILEP is suitable for quantitative plant proteomic analysis, where plants are grown in either (14)N- or (15)N-hydroponic media and pooled when the biological samples are collected for relative proteome quantitation. The pooled (14)N-/(15)N-protein extracts can be fractionated in any suitable way and digested with a protease for shotgun proteomics, using typically reverse phase liquid chromatography nanoelectrospray ionization tandem mass spectrometry (RPLC-nESI-MS/MS). Best results were obtained with a hybrid ion trap/FT-MS mass spectrometer, combining high mass accuracy and sensitivity for the MS data acquisition with speed and high-throughput MS/MS data acquisition, increasing the number of proteins identified and quantified and improving protein quantitation. Peak processing and picking from raw MS data files, protein identification, and quantitation were performed in a highly automated way using integrated MS data analysis software with minimum manual intervention, thus easing the analytical workflow. In this methodology paper, we describe how to grow Arabidopsis plants hydroponically for isotope labeling using (15)N salts and how to quantitate the resulting proteomes using a convenient workflow that does not require extensive bioinformatics skills.

  8. Spectral isotopic methods of determining nitrogen and carbon in plant specimens with laser volatization

    International Nuclear Information System (INIS)

    Lazeeva, G.S.

    1986-01-01

    Methods have been devised for the local determination of nitrogen and carbon isotope compositions in plant specimens, which provide separate and joint determination. Local laser evaporation has been combined with spectroscopic determination of the isotope compositions in the gas phase. A continuous-wave CO 2 laser is preferable for the local evaporation; the carbon isotope composition may be determined directly on the sum of the evaporation products, whereas nitrogen must first be separated as N 2 . Methods have also been developed for the local determination of total nitrogen and carbon in a sample with isotope dilution on the basis of laser evaporation. In order to eliminate systematic errors in determining total carbon in plant material, an evaporation method free from a rim has been devised. These methods have been used in determining isotope concentration profiles in plant specimens grown in experiments employing labeled nitrogen and carbon

  9. Subcellular SIMS imaging of isotopically labeled amino acids in cryogenically prepared cells

    International Nuclear Information System (INIS)

    Chandra, Subhash

    2004-01-01

    Ion microscopy is a potentially powerful technique for localization of isotopically labeled molecules. In this study, L-arginine and phenylalanine amino acids labeled with stable isotopes 13 C and 15 N were localized in cultured cells with the ion microscope at 500 nm spatial resolution. Cells were exposed to the labeled amino acids and cryogenically prepared. SIMS analyses were made in fractured freeze-dried cells. A dynamic distribution was observed from labeled arginine-treated LLC-PK 1 kidney cells at mass 28 ( 13 C 15 N) in negative secondaries, revealing cell-to-cell heterogeneity and preferential accumulation of the amino acid (or its metabolite) in the nucleus and nucleolus of some cells. The smaller nucleolus inside the nucleus was clearly resolved in SIMS images and confirmed by correlative light microscopy. The distribution of labeled phenylalanine contrasted with arginine as it was rather homogeneously distributed in T98G human glioblastoma cells. Images of 39 K, 23 Na and 40 Ca were also recorded to confirm the reliability of sample preparation and authenticity of the observed amino acid distributions. These observations indicate that SIMS techniques can provide a valuable technology for subcellular localization of nitrogen-containing molecules in proteomics since nitrogen does not have a radionuclide tracer isotope. Amino acids labeled with stable isotopes can be used as tracers for studying their transport and metabolism in distinct subcellular compartments with SIMS. Further studies of phenylalanine uptake in human glioblastoma cells may have special significance in boron neutron capture therapy (BNCT) as a boron analogue of phenylalanine, boronophenylalanine is a clinically approved compound for the treatment of brain tumors

  10. Subcellular SIMS imaging of isotopically labeled amino acids in cryogenically prepared cells

    Energy Technology Data Exchange (ETDEWEB)

    Chandra, Subhash

    2004-06-15

    Ion microscopy is a potentially powerful technique for localization of isotopically labeled molecules. In this study, L-arginine and phenylalanine amino acids labeled with stable isotopes {sup 13}C and {sup 15}N were localized in cultured cells with the ion microscope at 500 nm spatial resolution. Cells were exposed to the labeled amino acids and cryogenically prepared. SIMS analyses were made in fractured freeze-dried cells. A dynamic distribution was observed from labeled arginine-treated LLC-PK{sub 1} kidney cells at mass 28 ({sup 13}C{sup 15}N) in negative secondaries, revealing cell-to-cell heterogeneity and preferential accumulation of the amino acid (or its metabolite) in the nucleus and nucleolus of some cells. The smaller nucleolus inside the nucleus was clearly resolved in SIMS images and confirmed by correlative light microscopy. The distribution of labeled phenylalanine contrasted with arginine as it was rather homogeneously distributed in T98G human glioblastoma cells. Images of {sup 39}K, {sup 23}Na and {sup 40}Ca were also recorded to confirm the reliability of sample preparation and authenticity of the observed amino acid distributions. These observations indicate that SIMS techniques can provide a valuable technology for subcellular localization of nitrogen-containing molecules in proteomics since nitrogen does not have a radionuclide tracer isotope. Amino acids labeled with stable isotopes can be used as tracers for studying their transport and metabolism in distinct subcellular compartments with SIMS. Further studies of phenylalanine uptake in human glioblastoma cells may have special significance in boron neutron capture therapy (BNCT) as a boron analogue of phenylalanine, boronophenylalanine is a clinically approved compound for the treatment of brain tumors.

  11. A Breast Cell Atlas: Organelle analysis of the MDA-MB-231 cell line by density-gradient fractionation using isotopic marking and label-free analysis

    Directory of Open Access Journals (Sweden)

    Marianne Sandin

    2015-09-01

    Full Text Available Protein translocation between organelles in the cell is an important process that regulates many cellular functions. However, organelles can rarely be isolated to purity so several methods have been developed to analyse the fractions obtained by density gradient centrifugation. We present an analysis of the distribution of proteins amongst organelles in the human breast cell line, MDA-MB-231 using two approaches: an isotopic labelling and a label-free approach.

  12. An economic approach to efficient isotope labeling in insect cells using homemade {sup 15}N-, {sup 13}C- and {sup 2}H-labeled yeast extracts

    Energy Technology Data Exchange (ETDEWEB)

    Opitz, Christian; Isogai, Shin; Grzesiek, Stephan, E-mail: Stephan.Grzesiek@unibas.ch [University of Basel, Focal Area Structural Biology and Biophysics, Biozentrum (Switzerland)

    2015-07-15

    Heterologous expression of proteins in insect cells is frequently used for crystallographic structural studies due to the high yields even for challenging proteins requiring the eukaryotic protein processing capabilities of the host. However for NMR studies, the need for isotope labeling poses extreme challenges in eukaryotic hosts. Here, we describe a robust method to achieve uniform protein {sup 15}N and {sup 13}C labeling of up to 90 % in baculovirus-infected insect cells. The approach is based on the production of labeled yeast extract, which is subsequently supplemented to insect cell growth media. The method also allows deuteration at levels of >60 % without decrease in expression yield. The economic implementation of the labeling procedures into a standard structural biology laboratory environment is described in a step-by-step protocol. Applications are demonstrated for a variety of NMR experiments using the Abelson kinase domain, GFP, and the beta-1 adrenergic receptor as examples. Deuterated expression of the latter provides spectra of very high quality of a eukaryotic G-protein coupled receptor.

  13. An economic approach to efficient isotope labeling in insect cells using homemade 15N-, 13C- and 2H-labeled yeast extracts

    International Nuclear Information System (INIS)

    Opitz, Christian; Isogai, Shin; Grzesiek, Stephan

    2015-01-01

    Heterologous expression of proteins in insect cells is frequently used for crystallographic structural studies due to the high yields even for challenging proteins requiring the eukaryotic protein processing capabilities of the host. However for NMR studies, the need for isotope labeling poses extreme challenges in eukaryotic hosts. Here, we describe a robust method to achieve uniform protein 15 N and 13 C labeling of up to 90 % in baculovirus-infected insect cells. The approach is based on the production of labeled yeast extract, which is subsequently supplemented to insect cell growth media. The method also allows deuteration at levels of >60 % without decrease in expression yield. The economic implementation of the labeling procedures into a standard structural biology laboratory environment is described in a step-by-step protocol. Applications are demonstrated for a variety of NMR experiments using the Abelson kinase domain, GFP, and the beta-1 adrenergic receptor as examples. Deuterated expression of the latter provides spectra of very high quality of a eukaryotic G-protein coupled receptor

  14. Development of chemical isotope labeling liquid chromatography mass spectrometry for silkworm hemolymph metabolomics

    International Nuclear Information System (INIS)

    Shen, Weifeng; Han, Wei; Li, Yunong; Meng, Zhiqi; Cai, Leiming; Li, Liang

    2016-01-01

    -Methyl-D-aspartic and tyrosine were identified. The changes of these biomarkers were likely due to the disruption of the endocrine system of silkworm by DDT. This work illustrates that the method of CIL LC-MS is useful to generate quantitative submetabolome profiles from a small volume of silkworm hemolymph with much higher coverage than conventional LC-MS methods, thereby facilitating the discovery of potential metabolite biomarkers related to EDC or other chemical exposure. - Highlights: • A method was developed for profiling silkworm hemolymph metabolome with high coverage. • Dansylation isotope labeling LC-MS was used for amine/phenol submetabolome quantification. • Metabolomic changes in silkworm after DDT exposure were studied. • Among 2044 metabolites detected, 65 were positively identified and 1809 were putatively identified. • 33 positively identified metabolites showed significant changes in five groups of silkworms.

  15. Development of chemical isotope labeling liquid chromatography mass spectrometry for silkworm hemolymph metabolomics

    Energy Technology Data Exchange (ETDEWEB)

    Shen, Weifeng [Key Laboratory of Detection for Pesticide Residues, Ministry of Agriculture (China); Sericultural Research Institute, Zhejiang Academy of Agricultural Sciences, Hangzhou (China); Han, Wei; Li, Yunong [Department of Chemistry, University of Alberta, Edmonton, Alberta (Canada); Meng, Zhiqi [Sericultural Research Institute, Zhejiang Academy of Agricultural Sciences, Hangzhou (China); Cai, Leiming, E-mail: cailm@mail.zaas.ac.cn [Institute of Quality and Standard for Agro-products, Zhejiang Academy of Agricultural Sciences, Hangzhou (China); Li, Liang, E-mail: Liang.Li@ualberta.ca [Department of Chemistry, University of Alberta, Edmonton, Alberta (Canada)

    2016-10-26

    -Methyl-D-aspartic and tyrosine were identified. The changes of these biomarkers were likely due to the disruption of the endocrine system of silkworm by DDT. This work illustrates that the method of CIL LC-MS is useful to generate quantitative submetabolome profiles from a small volume of silkworm hemolymph with much higher coverage than conventional LC-MS methods, thereby facilitating the discovery of potential metabolite biomarkers related to EDC or other chemical exposure. - Highlights: • A method was developed for profiling silkworm hemolymph metabolome with high coverage. • Dansylation isotope labeling LC-MS was used for amine/phenol submetabolome quantification. • Metabolomic changes in silkworm after DDT exposure were studied. • Among 2044 metabolites detected, 65 were positively identified and 1809 were putatively identified. • 33 positively identified metabolites showed significant changes in five groups of silkworms.

  16. Mechanistic Insights into Catalytic Ethanol Steam Reforming Using Isotope-Labeled Reactants.

    Science.gov (United States)

    Crowley, Stephen; Castaldi, Marco J

    2016-08-26

    The low-temperature ethanol steam reforming (ESR) reaction mechanism over a supported Rh/Pt catalyst has been investigated using isotope-labeled EtOH and H2 O. Through strategic isotope labeling, all nonhydrogen atoms were distinct from one another, and allowed an unprecedented level of understanding of the dominant reaction pathways. All combinations of isotope- and non-isotope-labeled atoms were detected in the products, thus there are multiple pathways involved in H2 , CO, CO2 , CH4 , C2 H4 , and C2 H6 product formation. Both the recombination of C species on the surface of the catalyst and preservation of the C-C bond within ethanol are responsible for C2 product formation. Ethylene is not detected until conversion drops below 100 % at t=1.25 h. Also, quantitatively, 57 % of the observed ethylene is formed directly through ethanol dehydration. Finally there is clear evidence to show that oxygen in the SiO2 -ZrO2 support constitutes 10 % of the CO formed during the reaction. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. REDOR NMR of stable-isotope-labeled protein binding sites

    Energy Technology Data Exchange (ETDEWEB)

    Schaefer, J. [Washington Univ., St. Louis, MO (United States)

    1994-12-01

    Rotational-echo, double resonance (REDOR) NMR, a new analytical spectroscopic technique for solids spinning at the magic angle, has been developed over the last 5 years. REDOR provides a direct measure of heteronuclear dipolar coupling between isolated pairs of labeled nuclei. In a solid with a {sup 13}C-{sup 15}N labeled pair, for example, the {sup 13}C rotational echoes that form each rotor period following a{sup 1}H-{sup 13}C cross-polarization transfer can be prevented from reaching full intensity by insertion of a {sup 15}N {pi} pulse each half rotor period. The REDOR difference (the difference between a {sup 13}C NMR spectrum obtained under these conditions and one obtained with no {sup 15}N {pi} pulses) has a strong dependence on the {sup 13}C-{sup 15}N dipolar coupling, and hence, the {sup 13}C-{sup 15}N internuclear distance. REDOR is described as double-resonance even though three radio frequencies (typically {sup 1}H, {sup 13}C, and {sup 15}N) are used because the protons are removed from the important evolution part of the experiment by resonant decoupling. The dephasing of magnetization in REDOR arises from a local dipolar {sup 13}C-{sup 15}N field gradient and involves no polarization transfer. REDOR has no dependence on {sup 13}C or {sup 15}N chemical-shift tensors and does not require resolution of a {sup 13}C-{sup 15}N coupling in the chemical-shift dimension.

  18. A general procedure for isotopic (deuterium) labelling of non-steroidal antiinflammatory 2-arylpropionic acids

    International Nuclear Information System (INIS)

    Castell, J.V.; Martinez, L.A.; Universidad Politecnica de Valencia; Miranda, M.A.; Tarrega, Pilar

    1994-01-01

    Alkaline treatment of nonsteroidal antiinflammatory 2-arylpropionic acids in deuterium oxide led in all cases to isotopic exchange of the proton located at the α-position of the side chain. Monodeuteration was observed in the case of carprofen, ibuprofen, ketoprofen, fenoprofen, flurbiprofen and naproxen. Additional exchange of one or two protons of the heterocyclic ring occurred in indoprofen, suprofen and tiaprofenic acid. The isotopic labelling survived under the conditions required to perform in vitro photoallergic studies (photolysis in non-deuterated aqueous media). (Author)

  19. A general procedure for isotopic (deuterium) labelling of non-steroidal antiinflammatory 2-arylpropionic acids

    Energy Technology Data Exchange (ETDEWEB)

    Castell, J.V. (Valencia Univ. Hospital (Spain). Centro de Investigacion); Martinez, L.A. (Valencia Univ. Hospital (Spain). Centro de Investigacion Universidad Politecnica de Valencia (Spain). Dept. de Quimica); Miranda, M.A.; Tarrega, Pilar (Universidad Politecnica de Valencia (Spain). Dept. de Quimica)

    1994-01-01

    Alkaline treatment of nonsteroidal antiinflammatory 2-arylpropionic acids in deuterium oxide led in all cases to isotopic exchange of the proton located at the [alpha]-position of the side chain. Monodeuteration was observed in the case of carprofen, ibuprofen, ketoprofen, fenoprofen, flurbiprofen and naproxen. Additional exchange of one or two protons of the heterocyclic ring occurred in indoprofen, suprofen and tiaprofenic acid. The isotopic labelling survived under the conditions required to perform in vitro photoallergic studies (photolysis in non-deuterated aqueous media). (Author).

  20. Quantitative amino acid profiling and stable isotopically labeled amino acid tracer enrichment used for in vivo human systemic and tissue kinetics measurements

    DEFF Research Database (Denmark)

    Bornø, Andreas; van Hall, Gerrit

    2014-01-01

    An important area within clinical functional metabolomics is in vivo amino acid metabolism and protein turnover measurements for which accurate amino acid concentrations and stable isotopically labeled amino acid enrichments are mandatory not the least when tissue metabolomics is determined....... The present study describes a new sensitive liquid chromatography tandem mass-spectrometry method quantifying 20 amino acids and their tracer(s) ([ring-(13)C6]/D5Phenylalanine) in human plasma and skeletal muscle specimens. Before analysis amino acids were extracted and purified via deprotonization....../ion exchange, derivatized using a phenylisothiocyanate reagent and each amino acid was quantitated with its own stable isotopically labeled internal standard (uniformly labeled-(13)C/(15)N). The method was validated according to general recommendations for chromatographic analytical methods. The calibration...

  1. The doubly-labelled water method for measuring energy expenditure. Technical recommendations for use in humans

    International Nuclear Information System (INIS)

    Prentice, A.M.

    1990-01-01

    The doubly-labelled water method using stable isotopes of hydrogen and oxygen, is rapidly becoming established as an important new tool for investigating energy metabolism. It is the first genuinely non-invasive method for measuring energy expenditure in free-living people, providing estimates of habitual expenditure over a time period of 10-20 days. The accuracy and precision of these estimates should be superior to those obtained by traditional factorial methods. The DLW method involves several assumptions about the behaviour of the isotopes, the body water pool and the exchange rates within that pool in the labelled animal. These assumptions are: (1) The volume of the body water pool remains constant throughout the measurement period. (2) The rates of water influx, and water and CO 2 efflux are constant throughout the measurement period. (3) The isotopes label only the H 2 O and CO 2 in the body. (4) The isotopes leave the body only in the form of H 2 O and CO 2 . (5) The concentrations of the isotopes in H 2 O and CO 2 leaving the body are the same as those in body water at that time (i.e. there is no isotopic fractionation). (6) No H 2 O or CO 2 that has left the body re-enters the body. (7) The natural abundance, or ''background'' levels of the isotopes remain constant during the measurement interval. The recommendations presented in this document are based on a variety of criteria including: (a) which procedure among several is theoretically correct in a given application; (b) which procedure is simplest and least prone to methodological errors; and (c) which procedure yields the lowest error in validation studies. Refs, figs and tabs

  2. Chemical Ligation and Isotope Labeling to Locate Dynamic Effects during Catalysis by Dihydrofolate Reductase.

    Science.gov (United States)

    Luk, Louis Y P; Ruiz-Pernía, J Javier; Adesina, Aduragbemi S; Loveridge, E Joel; Tuñón, Iñaki; Moliner, Vincent; Allemann, Rudolf K

    2015-07-27

    Chemical ligation has been used to alter motions in specific regions of dihydrofolate reductase from E. coli and to investigate the effects of localized motional changes on enzyme catalysis. Two isotopic hybrids were prepared; one with the mobile N-terminal segment containing heavy isotopes ((2) H, (13) C, (15) N) and the remainder of the protein with natural isotopic abundance, and the other one with only the C-terminal segment isotopically labeled. Kinetic investigations indicated that isotopic substitution of the N-terminal segment affected only a physical step of catalysis, whereas the enzyme chemistry was affected by protein motions from the C-terminal segment. QM/MM studies support the idea that dynamic effects on catalysis mostly originate from the C-terminal segment. The use of isotope hybrids provides insights into the microscopic mechanism of dynamic coupling, which is difficult to obtain with other studies, and helps define the dynamic networks of intramolecular interactions central to enzyme catalysis. © 2015 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.

  3. Development of high-performance chemical isotope labeling LC-MS for profiling the human fecal metabolome.

    Science.gov (United States)

    Xu, Wei; Chen, Deying; Wang, Nan; Zhang, Ting; Zhou, Ruokun; Huan, Tao; Lu, Yingfeng; Su, Xiaoling; Xie, Qing; Li, Liang; Li, Lanjuan

    2015-01-20

    Human fecal samples contain endogenous human metabolites, gut microbiota metabolites, and other compounds. Profiling the fecal metabolome can produce metabolic information that may be used not only for disease biomarker discovery, but also for providing an insight about the relationship of the gut microbiome and human health. In this work, we report a chemical isotope labeling liquid chromatography-mass spectrometry (LC-MS) method for comprehensive and quantitative analysis of the amine- and phenol-containing metabolites in fecal samples. Differential (13)C2/(12)C2-dansyl labeling of the amines and phenols was used to improve LC separation efficiency and MS detection sensitivity. Water, methanol, and acetonitrile were examined as an extraction solvent, and a sequential water-acetonitrile extraction method was found to be optimal. A step-gradient LC-UV setup and a fast LC-MS method were evaluated for measuring the total concentration of dansyl labeled metabolites that could be used for normalizing the sample amounts of individual samples for quantitative metabolomics. Knowing the total concentration was also useful for optimizing the sample injection amount into LC-MS to maximize the number of metabolites detectable while avoiding sample overloading. For the first time, dansylation isotope labeling LC-MS was performed in a simple time-of-flight mass spectrometer, instead of high-end equipment, demonstrating the feasibility of using a low-cost instrument for chemical isotope labeling metabolomics. The developed method was applied for profiling the amine/phenol submetabolome of fecal samples collected from three families. An average of 1785 peak pairs or putative metabolites were found from a 30 min LC-MS run. From 243 LC-MS runs of all the fecal samples, a total of 6200 peak pairs were detected. Among them, 67 could be positively identified based on the mass and retention time match to a dansyl standard library, while 581 and 3197 peak pairs could be putatively

  4. The synthesis of tritium, carbon-14 and stable isotope labelled selective estrogen receptor degraders.

    Science.gov (United States)

    Bragg, Ryan A; Bushby, Nick; Ericsson, Cecilia; Kingston, Lee P; Ji, Hailong; Elmore, Charles S

    2016-09-01

    As part of a Medicinal Chemistry program aimed at developing an orally bioavailable selective estrogen receptor degrader, a number of tritium, carbon-14, and stable isotope labelled (E)-3-[4-(2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indol-1-yl)phenyl]prop-2-enoic acids were required. This paper discusses 5 synthetic approaches to this compound class. Copyright © 2016 John Wiley & Sons, Ltd.

  5. Isotopic ratio method for determining uranium contamination

    International Nuclear Information System (INIS)

    Miles, R.E.; Sieben, A.K.

    1994-01-01

    The presence of high concentrations of uranium in the subsurface can be attributed either to contamination from uranium processing activities or to naturally occurring uranium. A mathematical method has been employed to evaluate the isotope ratios from subsurface soils at the Rocky Flats Nuclear Weapons Plant (RFP) and demonstrates conclusively that the soil contains uranium from a natural source and has not been contaminated with enriched uranium resulting from RFP releases. This paper describes the method used in this determination which has widespread application in site characterizations and can be adapted to other radioisotopes used in manufacturing industries. The determination of radioisotope source can lead to a reduction of the remediation effort

  6. Determination of symbiotic nitrogen fixation by labelling the soil atmosphere with sup(15)N sub(2) at low isotope enrichment

    International Nuclear Information System (INIS)

    Trivelin, P.C.O.

    1982-01-01

    A direct method to determine the total symbiotic nitrogen fixation during the leguminous plants cycles has been, developed, by labelling the soil atmosphere with sup(15)N sub(2) at low isotope enrichment, of about 1 atom % excess. The soil explored by the root system of leguminous plants was confined by means of a chamber in the field and by sealed pots in greenhouse experiments in order to maintain the soil air labelled with sup(15)N sub(2). The average sup(15)N concentration in the soil atmosphere, necessary to calculate dinitrogen fixation, was obtained by integration of the exponential functions of isotope dilution. Those functions were obtained by periodic sampling and analysis of the N sub(2) in the soil atmosphere. The field experiment with labelled atmosphere was carried out from the 22 sup(nd) to the 31 sup(st) day of the bean crop cycle and 5.5 mg N/plant (24% of total plant N) was derived from fixation. In pot experiments, under greenhouse conditions, integrated determination of fixation was made in Phaseolus beans (from the 19 sup(th) to the 67 sup(th) day from planting) and in soybeans (from the 24 sup(th) to the 70 sup(th) day from planting). The soil atmosphere was labelled with sup(15)N sub(2) in both cases. Average fixation obtained for Phaseolus beans was 80 mg N/plant (65% of total plant N) and for soybeans 265 mg N/plant (71% of total plant N). Evaluation of the basic concept of the isotope dilution method to determine nitrogen fixation in pots experiments, as proposed by Fried and Middelboe (1977) has also been made in the present paper. Simultaneous determinations of fixation in soybeans, using the isotope dilution method of Fried and Middelboe, natural variation of the sup(15)N/ sup(14)N ratios, and total-N differences, indicated the same results for pot experiments, harvested at the end of the plant cycle. (author)

  7. Evaluation of the impact of matrix effect on quantification of pesticides in foods by gas chromatography-mass spectrometry using isotope-labeled internal standards.

    Science.gov (United States)

    Yarita, Takashi; Aoyagi, Yoshie; Otake, Takamitsu

    2015-05-29

    The impact of the matrix effect in GC-MS quantification of pesticides in food using the corresponding isotope-labeled internal standards was evaluated. A spike-and-recovery study of nine target pesticides was first conducted using paste samples of corn, green soybean, carrot, and pumpkin. The observed analytical values using isotope-labeled internal standards were more accurate for most target pesticides than that obtained using the external calibration method, but were still biased from the spiked concentrations when a matrix-free calibration solution was used for calibration. The respective calibration curves for each target pesticide were also prepared using matrix-free calibration solutions and matrix-matched calibration solutions with blank soybean extract. The intensity ratio of the peaks of most target pesticides to that of the corresponding isotope-labeled internal standards was influenced by the presence of the matrix in the calibration solution; therefore, the observed slope varied. The ratio was also influenced by the type of injection method (splitless or on-column). These results indicated that matrix-matching of the calibration solution is required for very accurate quantification, even if isotope-labeled internal standards were used for calibration. Copyright © 2015 Elsevier B.V. All rights reserved.

  8. On structural identifiability analysis of the cascaded linear dynamic systems in isotopically non-stationary 13C labelling experiments.

    Science.gov (United States)

    Lin, Weilu; Wang, Zejian; Huang, Mingzhi; Zhuang, Yingping; Zhang, Siliang

    2018-06-01

    The isotopically non-stationary 13C labelling experiments, as an emerging experimental technique, can estimate the intracellular fluxes of the cell culture under an isotopic transient period. However, to the best of our knowledge, the issue of the structural identifiability analysis of non-stationary isotope experiments is not well addressed in the literature. In this work, the local structural identifiability analysis for non-stationary cumomer balance equations is conducted based on the Taylor series approach. The numerical rank of the Jacobian matrices of the finite extended time derivatives of the measured fractions with respect to the free parameters is taken as the criterion. It turns out that only one single time point is necessary to achieve the structural identifiability analysis of the cascaded linear dynamic system of non-stationary isotope experiments. The equivalence between the local structural identifiability of the cascaded linear dynamic systems and the local optimum condition of the nonlinear least squares problem is elucidated in the work. Optimal measurements sets can then be determined for the metabolic network. Two simulated metabolic networks are adopted to demonstrate the utility of the proposed method. Copyright © 2018 Elsevier Inc. All rights reserved.

  9. Liquid–liquid extraction combined with differential isotope dimethylaminophenacyl labeling for improved metabolomic profiling of organic acids

    International Nuclear Information System (INIS)

    Peng, Jun; Li, Liang

    2013-01-01

    Graphical abstract: -- Highlights: •An improved method for profiling the carboxylic acid sub-metabolome is reported. •Liquid–liquid extraction was used for separating the organic acids from the amines. • 12 C/ 13 C-p-dimethylaminophenacyl (DmPA) labeling of the organic acids was carried out on the extract. •Detection interference by amines and labeling efficiency reduction by water were reduced. •About 2500 12 C/ 13 C-peak pairs or putative metabolites could be detected from 20 μL of human urine. -- Abstract: A large fraction of the known human metabolome belong to organic acids. However, comprehensive profiling of the organic acid sub-metabolome is a major analytical challenge. In this work, we report an improved method for detecting organic acid metabolites. This method is based on the use of liquid–liquid extraction (LLE) to selectively extract the organic acids, followed by using differential isotope p-dimethylaminophenacyl (DmPA) labeling of the acid metabolites. The 12 C-/ 13 C-labeled samples are analyzed by liquid chromatography Fourier-transform ion cyclotron resonance mass spectrometry (LC–FTICR–MS). It is shown that this LLE DmPA labeling method offers superior performance over the method of direct DmPA labeling of biofluids such as human urine. LLE of organic acids reduces the interference of amine-containing metabolites that may also react with DmPA. It can also remove water in a biofluid that can reduce the labeling efficiency. Using human urine as an example, it is demonstrated that about 2500 peak pairs or putative metabolites could be detected in a 30-min gradient LC–MS run, which is about 3 times more than that detected in a sample prepared using direct DmPA labeling. About 95% of the 1000 or so matched metabolites to the Human Metabolome Database (HMDB) are organic acids. It is further shown that this method can be used to handle as small as 10 μL of urine. We believe that this method opens the possibility of generating a

  10. Liquid–liquid extraction combined with differential isotope dimethylaminophenacyl labeling for improved metabolomic profiling of organic acids

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Jun; Li, Liang, E-mail: Liang.Li@ualberta.ca

    2013-11-25

    Graphical abstract: -- Highlights: •An improved method for profiling the carboxylic acid sub-metabolome is reported. •Liquid–liquid extraction was used for separating the organic acids from the amines. •{sup 12}C/{sup 13}C-p-dimethylaminophenacyl (DmPA) labeling of the organic acids was carried out on the extract. •Detection interference by amines and labeling efficiency reduction by water were reduced. •About 2500 {sup 12}C/{sup 13}C-peak pairs or putative metabolites could be detected from 20 μL of human urine. -- Abstract: A large fraction of the known human metabolome belong to organic acids. However, comprehensive profiling of the organic acid sub-metabolome is a major analytical challenge. In this work, we report an improved method for detecting organic acid metabolites. This method is based on the use of liquid–liquid extraction (LLE) to selectively extract the organic acids, followed by using differential isotope p-dimethylaminophenacyl (DmPA) labeling of the acid metabolites. The {sup 12}C-/{sup 13}C-labeled samples are analyzed by liquid chromatography Fourier-transform ion cyclotron resonance mass spectrometry (LC–FTICR–MS). It is shown that this LLE DmPA labeling method offers superior performance over the method of direct DmPA labeling of biofluids such as human urine. LLE of organic acids reduces the interference of amine-containing metabolites that may also react with DmPA. It can also remove water in a biofluid that can reduce the labeling efficiency. Using human urine as an example, it is demonstrated that about 2500 peak pairs or putative metabolites could be detected in a 30-min gradient LC–MS run, which is about 3 times more than that detected in a sample prepared using direct DmPA labeling. About 95% of the 1000 or so matched metabolites to the Human Metabolome Database (HMDB) are organic acids. It is further shown that this method can be used to handle as small as 10 μL of urine. We believe that this method opens the

  11. Sulfur Isotope Exchange between S-35 Labeled Inorganic Sulfur-Compounds in Anoxic Marine-Sediments

    DEFF Research Database (Denmark)

    FOSSING, H.; THODEANDERSEN, S.; JØRGENSEN, BB

    1992-01-01

    Isotope exchange reactions between S-35-labeled sulfur compounds were studied in anoxic estuarine sediment slurries at 21-degrees-C and pH 7.4-7.7. Two experiments labeled with radioactive elemental sulfur (S-35-degrees) and one labeled with radioactive sulfate ((SO42-)-S-35) were performed as time......% of the total S-35 was recovered in the SIGMA-HS- pool in less than 1.5 h. With no detectable SIGMA-HS- (less than 1-mu-M) in the slurry, 58% of the total S-35 was observed in the pyrite pool within 1.5 h. The FeS pool received up to 31% of all S-35 added. The rapid S-35 incorporation from S-35-degrees...... into SIGMA-HS- and FeS pools was explained by isotope exchange reactions. In contrast, there was evidence that the radioactivity observed in the 'pyrite pool' was caused by adhesion of the added S-35-degrees to the FeS2 grains. In all S-35-degrees-labeled experiments we also observed oxidation...

  12. Metabolic Flux Analysis in Isotope Labeling Experiments Using the Adjoint Approach.

    Science.gov (United States)

    Mottelet, Stephane; Gaullier, Gil; Sadaka, Georges

    2017-01-01

    Comprehension of metabolic pathways is considerably enhanced by metabolic flux analysis (MFA-ILE) in isotope labeling experiments. The balance equations are given by hundreds of algebraic (stationary MFA) or ordinary differential equations (nonstationary MFA), and reducing the number of operations is therefore a crucial part of reducing the computation cost. The main bottleneck for deterministic algorithms is the computation of derivatives, particularly for nonstationary MFA. In this article, we explain how the overall identification process may be speeded up by using the adjoint approach to compute the gradient of the residual sum of squares. The proposed approach shows significant improvements in terms of complexity and computation time when it is compared with the usual (direct) approach. Numerical results are obtained for the central metabolic pathways of Escherichia coli and are validated against reference software in the stationary case. The methods and algorithms described in this paper are included in the sysmetab software package distributed under an Open Source license at http://forge.scilab.org/index.php/p/sysmetab/.

  13. Classification-based quantitative analysis of stable isotope labeling by amino acids in cell culture (SILAC) data.

    Science.gov (United States)

    Kim, Seongho; Carruthers, Nicholas; Lee, Joohyoung; Chinni, Sreenivasa; Stemmer, Paul

    2016-12-01

    Stable isotope labeling by amino acids in cell culture (SILAC) is a practical and powerful approach for quantitative proteomic analysis. A key advantage of SILAC is the ability to simultaneously detect the isotopically labeled peptides in a single instrument run and so guarantee relative quantitation for a large number of peptides without introducing any variation caused by separate experiment. However, there are a few approaches available to assessing protein ratios and none of the existing algorithms pays considerable attention to the proteins having only one peptide hit. We introduce new quantitative approaches to dealing with SILAC protein-level summary using classification-based methodologies, such as Gaussian mixture models with EM algorithms and its Bayesian approach as well as K-means clustering. In addition, a new approach is developed using Gaussian mixture model and a stochastic, metaheuristic global optimization algorithm, particle swarm optimization (PSO), to avoid either a premature convergence or being stuck in a local optimum. Our simulation studies show that the newly developed PSO-based method performs the best among others in terms of F1 score and the proposed methods further demonstrate the ability of detecting potential markers through real SILAC experimental data. No matter how many peptide hits the protein has, the developed approach can be applicable, rescuing many proteins doomed to removal. Furthermore, no additional correction for multiple comparisons is necessary for the developed methods, enabling direct interpretation of the analysis outcomes. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  14. Validation of a dual-isotope plasma ratio method for measurement of cholesterol absorption in rats

    International Nuclear Information System (INIS)

    Zilversmit, D.B.; Hughes, L.B.

    1974-01-01

    Several methods for measuring cholesterol absorption in the rat have been compared. After administration of an oral dose of labeled cholesterol ( 14 C or 3 H) and an intravenous dose of colloidal labeled cholesterol ( 3 H or 14 C) the ratio of the two labels in plasma or whole blood 48 hr or more after dosing compared closely to the ratio of areas under the respective specific activity-time curves. The area ratio method is independent of a time lag between the appearance of oral and intravenous label in the bloodstream. Both measures of cholesterol absorption agree fairly well with a method based on measuring the unabsorbed dietary cholesterol in a pooled fecal sample. The plasma isotope ratio method gave more reproducible results than the fecal collection method when the measurement was repeated in the same animals 5 days after the first measurement. Cholesterol absorption was overestimated by the use of Tween 20-solubilized labeled cholesterol for the intravenous dose. The plasma disappearance curves of injected labeled colloidal cholesterol and cholesterol-labeled chylomicrons infused intravenously over a 3.5-h period in the same animal coincided within experimental error from the first day until 75 days after injection. The plasma isotope ratio method for cholesterol absorption gave the same results in rats practicing coprophagy as in those in which this practice was prevented. The addition of sulfaguanidine to the diet lowered cholesterol absorption as measured by the plasma isotope ratio to the same degree as that measured by the fecal collection method. (U.S.)

  15. Cooperation of CMEA member states in the field of the manufacture and use of stable isotopes and compounds thus labelled

    International Nuclear Information System (INIS)

    Ertel, G.; Ewald, G.

    1977-01-01

    The contribution presents a survey of scientific-technical cooperation of CMEA member states in the field of stable isotopes, it deals with the specialization of stable isotope production and compounds thus labelled, and gives the prospects for further development of this cooperation. (HK) [de

  16. Metabolic flux analysis of the phenylpropanoid pathway in wound-healing potato tuber tissue using stable isotope-labeled tracer and LC-MS spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Matsuda, Fumio; Morino, Keiko; Miyashita, Masahiro; Miyagawa, Hisashi [Kyoto Univ. (Japan). Department of Agriculture

    2003-05-01

    The metabolic flux of two phenylpropanoid metabolites, N-p-coumaroyloctopamine (p-CO) and chlorogenic acid (CGA), in the wound-healing potato tuber tissue was quantitatively analyzed by a newly developed method based upon the tracer experiment using stable isotope-labeled compounds and LC-MS. Tuber disks were treated with aqueous solution of L-phenyl-d{sub 5}-alanine, and the change in the ratio of stable isotope-labeled compound to non-labeled (isotope abundance) was monitored for p-CO and CGA in the tissue extract by LC-MS. The time-dependent change in the isotope abundance of each metabolite was fitted to an equation that was derived from the formation and conversion kinetics of each compound. Good correlations were obtained between the observed and calculated isotope abundances for both p-CO and CGA. The rates of p-CO formation and conversion (i.e. fluxes) were 1.15 and 0.96 nmol (g FW){sup -1}h{sup -1}, respectively, and for CGA, the rates 4.63 and 0.42 nmol (g FW){sup -1}h{sup -1}, respectively. This analysis enabled a direct comparison of the biosynthetic activity between these two compounds. (author)

  17. Stable isotope labeled n-alkanes to assess digesta passage kinetics through the digestive tract of ruminants

    NARCIS (Netherlands)

    Warner, D.; Ferreira, L.M.M.; Breuer, M.J.H.; Dijkstra, J.; Pellikaan, W.F.

    2013-01-01

    We describe the use of carbon stable isotope (13C) labeled n-alkanes as a potential internal tracer to assess passage kinetics of ingested nutrients in ruminants. Plant cuticular n-alkanes originating from intrinsically 13C labeled ryegrass plants were pulse dosed intraruminally in four

  18. Segmental isotope labeling of proteins for NMR structural study using a protein S tag for higher expression and solubility

    International Nuclear Information System (INIS)

    Kobayashi, Hiroshi; Swapna, G. V. T.; Wu, Kuen-Phon; Afinogenova, Yuliya; Conover, Kenith; Mao, Binchen; Montelione, Gaetano T.; Inouye, Masayori

    2012-01-01

    A common obstacle to NMR studies of proteins is sample preparation. In many cases, proteins targeted for NMR studies are poorly expressed and/or expressed in insoluble forms. Here, we describe a novel approach to overcome these problems. In the protein S tag-intein (PSTI) technology, two tandem 92-residue N-terminal domains of protein S (PrS 2 ) from Myxococcus xanthus is fused at the N-terminal end of a protein to enhance its expression and solubility. Using intein technology, the isotope-labeled PrS 2 -tag is replaced with non-isotope labeled PrS 2 -tag, silencing the NMR signals from PrS 2 -tag in isotope-filtered 1 H-detected NMR experiments. This method was applied to the E. coli ribosome binding factor A (RbfA), which aggregates and precipitates in the absence of a solubilization tag unless the C-terminal 25-residue segment is deleted (RbfAΔ25). Using the PrS 2 -tag, full-length well-behaved RbfA samples could be successfully prepared for NMR studies. PrS 2 (non-labeled)-tagged RbfA (isotope-labeled) was produced with the use of the intein approach. The well-resolved TROSY-HSQC spectrum of full-length PrS 2 -tagged RbfA superimposes with the TROSY-HSQC spectrum of RbfAΔ25, indicating that PrS 2 -tag does not affect the structure of the protein to which it is fused. Using a smaller PrS-tag, consisting of a single N-terminal domain of protein S, triple resonance experiments were performed, and most of the backbone 1 H, 15 N and 13 C resonance assignments for full-length E. coli RbfA were determined. Analysis of these chemical shift data with the Chemical Shift Index and heteronuclear 1 H– 15 N NOE measurements reveal the dynamic nature of the C-terminal segment of the full-length RbfA protein, which could not be inferred using the truncated RbfAΔ25 construct. CS-Rosetta calculations also demonstrate that the core structure of full-length RbfA is similar to that of the RbfAΔ25 construct.

  19. Isotope separation in plasma by ion-cyclotron resonance method

    International Nuclear Information System (INIS)

    Dubinov, A.E.; Kornilova, I.Yu.; Selemir, V.D.

    2001-01-01

    Contemporary state of investigation on isotope separation in plasma using selective ion-cyclotron resonance (ICR) heating is considered. The main attention is paid to necessary conditions of heating selectivity, plasma creation methods in isotope ICR-separation facilities, selection of antenna systems for heating, and principles of more-heated component selection. Experimental results obtained at different isotope mixtures separation are presented [ru

  20. Separation of the mercury isotopes by the indirect photochemical method

    International Nuclear Information System (INIS)

    Botter nee Bergheaud, F.; Scaringella nee Desnoyer, M.; Wacongne, M.

    1976-01-01

    A method of photochemical separation of the mercury isotopes by the so-called indirect route in which a gas stream of oxygen and butadiene containing a mixture of mercury isotopes is passed through one or a number of vessels placed in series. The gas stream is irradiated by a lamp containing mercury which is depleted in one or a number of the isotopes and said isotopes are recovered in a trap placed downstream of the vessel or vessels

  1. Recycling of an amino acid label with prolonged isotope infusion: Implications for kinetic studies

    International Nuclear Information System (INIS)

    Schwenk, W.F.; Tsalikian, E.; Beaufrere, B.; Haymond, M.W.

    1985-01-01

    To investigate whether recycling of a labeled amino acid would occur after 24 h of infusion, two groups of normal volunteers were infused with [ 3 H]leucine and alpha-[ 14 C]-ketoisocaproate for 4 h and [ 2 H 3 ]leucine for either 4 or 24 h (groups I and II, respectively). Entry of [ 2 H 3 ]leucine at steady state into the plasma space was indistinguishable from its infusion rate for group I but 30% higher (P less than 0.001) than this rate for group II, demonstrating significant recycling of label. After discontinuation of the infusions, isotope disappearance from the plasma space was followed for 2 h. The 3 H and 14 C decay data for both groups suggest that plasma leucine and alpha- ketoisocaproate are derived from a single intracellular pool in the postabsorptive state. In group I, the 3 H and 2 H labels decayed identically; whereas, in group II, the decay of [ 2 H 3 ]-leucine and alpha- [ 2 H 3 ]ketoisocaproate was slower (P less than 0.01) than the decay of [ 3 H]leucine and alpha-[ 3 H]ketoisocaproate, confirming re-entry of label after a 24-h infusion. Therefore kinetic values calculated from models assuming no recycling of labeled amino acids are most likely not quantitative and must be interpreted with care when flux does not change or decreases

  2. Selectively dispersed isotope labeling for protein structure determination by magic angle spinning NMR

    Energy Technology Data Exchange (ETDEWEB)

    Eddy, Matthew T. [Massachusetts Institute of Technology, Department of Chemistry (United States); Belenky, Marina [Brandeis University, Department of Chemistry (United States); Sivertsen, Astrid C. [Massachusetts Institute of Technology, Francis Bitter Magnet Laboratory (United States); Griffin, Robert G. [Massachusetts Institute of Technology, Department of Chemistry (United States); Herzfeld, Judith, E-mail: herzfeld@brandeis.edu [Brandeis University, Department of Chemistry (United States)

    2013-10-15

    The power of nuclear magnetic resonance spectroscopy derives from its site-specific access to chemical, structural and dynamic information. However, the corresponding multiplicity of interactions can be difficult to tease apart. Complimentary approaches involve spectral editing on the one hand and selective isotope substitution on the other. Here we present a new 'redox' approach to the latter: acetate is chosen as the sole carbon source for the extreme oxidation numbers of its two carbons. Consistent with conventional anabolic pathways for the amino acids, [1-{sup 13}C] acetate does not label {alpha} carbons, labels other aliphatic carbons and the aromatic carbons very selectively, and labels the carboxyl carbons heavily. The benefits of this labeling scheme are exemplified by magic angle spinning spectra of microcrystalline immunoglobulin binding protein G (GB1): the elimination of most J-couplings and one- and two-bond dipolar couplings provides narrow signals and long-range, intra- and inter-residue, recoupling essential for distance constraints. Inverse redox labeling, from [2-{sup 13}C] acetate, is also expected to be useful: although it retains one-bond couplings in the sidechains, the removal of CA-CO coupling in the backbone should improve the resolution of NCACX spectra.

  3. Automatic isotope gas analysis of tritium labelled organic materials Pt. 1

    International Nuclear Information System (INIS)

    Gacs, I.; Mlinko, S.

    1978-01-01

    A new automatic procedure developed to convert tritium in HTO hydrogen for subsequent on-line gas counting is described. The water containing tritium is introduced into a column prepared from molecular sieve-5A and heated to 550 deg C. The tritium is transferred by isotopic exchange into hydrogen flowing through the column. The radioactive gas is led into an internal detector for radioactivity measurement. The procedure is free of memory effects, provides quantitative recovery with analytical reproducibility better than 0.5% rel. at a preset number of counts. The experimental and analytical results indicate that isotopic exchange between HTO and hydrogen over a column prepared from alumina or molecular sieve-5A can be successfully applied for the quantitative transfer of tritium from HTO into hydrogen for on-line gas countinq. This provides an analytical procedure for the automatic determination of tritium in water with an analytical reproducibility better than 0.5% rel. The exchange process will also be suitable for rapid tritium transfer from water formed during the decomposition of tritium-labelled organic compounds or biological materials. The application of the procedure in automatic isotope gas analysis of organic materials labelled with tritium will be described in subsequent papers (Parts II and III). (T.G.)

  4. Development of uniformly stable isotope labeling system in higher plants for hetero-nuclear NMR experiments in vitro and in vivo

    International Nuclear Information System (INIS)

    Kikuchi, J.

    2005-01-01

    Full text: Novel methods for measurement of living systems are making new breakthroughs in life science. In the era of the metabolome (analysis of all measurable metabolites), a MS-based approach is considered to be the major technology, whereas a NMR-based method is recognized as minor technology due to its low sensitivity. Therefore, my laboratory is currently focusing to develop novel methodologies for an NMR-based metabolomics. This will be achieved by uniform stable isotope labeling of higher plants allowing application of multi-dimensional NMR experiments used in protein structure determination. Using these novel methods, I will analyze the dynamic molecular networks inside tissues. Especially, use of stable isotope labeling methods has enormous advantage for discrimination of incorporated or de novo synthesized compounds. Furthermore, potentiality of in vivo-NMR metabolomics will be discussed in the conference. (author)

  5. Double isotopic method using dansyl chloride for the determination of GABA in rat C6 astrocytoma cell cultures

    Energy Technology Data Exchange (ETDEWEB)

    Kohl, R.L.; Quay, W.B.; Perez-Polo, J.R.

    1986-01-01

    Methods are described for the quantitative measurement of GABA in culture. The method can be adapted to any amino acid or dansyl-chloride-reactive species. The sensitivity and selectivity of the procedure result from the double isotopic design in which (/sup 14/C)-labeled internal standard was added to the samples before reaction with (3M)-labeled dansyl chloride. Values obtained by ion-exchange amino acid analysis of cultures agree closely with the values obtained by the double isotopic method. This method is sensitive enough to measure GABA intracellularly and the condition medium.

  6. Systematic NMR Analysis of Stable Isotope Labeled Metabolite Mixtures in Plant and Animal Systems: Coarse Grained Views of Metabolic Pathways

    Science.gov (United States)

    Chikayama, Eisuke; Suto, Michitaka; Nishihara, Takashi; Shinozaki, Kazuo; Hirayama, Takashi; Kikuchi, Jun

    2008-01-01

    Background Metabolic phenotyping has become an important ‘bird's-eye-view’ technology which can be applied to higher organisms, such as model plant and animal systems in the post-genomics and proteomics era. Although genotyping technology has expanded greatly over the past decade, metabolic phenotyping has languished due to the difficulty of ‘top-down’ chemical analyses. Here, we describe a systematic NMR methodology for stable isotope-labeling and analysis of metabolite mixtures in plant and animal systems. Methodology/Principal Findings The analysis method includes a stable isotope labeling technique for use in living organisms; a systematic method for simultaneously identifying a large number of metabolites by using a newly developed HSQC-based metabolite chemical shift database combined with heteronuclear multidimensional NMR spectroscopy; Principal Components Analysis; and a visualization method using a coarse-grained overview of the metabolic system. The database contains more than 1000 1H and 13C chemical shifts corresponding to 142 metabolites measured under identical physicochemical conditions. Using the stable isotope labeling technique in Arabidopsis T87 cultured cells and Bombyx mori, we systematically detected >450 HSQC peaks in each 13C-HSQC spectrum derived from model plant, Arabidopsis T87 cultured cells and the invertebrate animal model Bombyx mori. Furthermore, for the first time, efficient 13C labeling has allowed reliable signal assignment using analytical separation techniques such as 3D HCCH-COSY spectra in higher organism extracts. Conclusions/Significance Overall physiological changes could be detected and categorized in relation to a critical developmental phase change in B. mori by coarse-grained representations in which the organization of metabolic pathways related to a specific developmental phase was visualized on the basis of constituent changes of 56 identified metabolites. Based on the observed intensities of 13C atoms of

  7. Method for the enrichment of isotopes

    International Nuclear Information System (INIS)

    Schuster, E.; Kersting, A.; Gebauhr, W.

    1976-01-01

    A method for the enrichment of isotopes, especially of uranium compounds, is described, working according to the principle of selective excitation by laser beams. As the starting compound, UF 6 is mentioned; the reaction partness are atomic gases (hydrogen, oxygen, nitrogen) or radicals (CH 3 , CHO). According to the invention, the gas mixture flows through the reaction space with a velocity so large that in leaving it, recombination of the reaction partner will already take place again. In this way, competing thermal reactions will be avoided and energy transfer fo excited 235 U molecules to non-excited 238 U molecules is eliminated to a large extent. A suitable equipment for implementing the method is described. (UWI) [de

  8. Quality Assurance of technitium-labelled radiopharmaceuticals in the Radiation and Isotopes Centre of Khartoum (RICK)

    International Nuclear Information System (INIS)

    Adlan, A.A.

    2005-09-01

    This descriptive, exploratory study was conducted in the nuclear medicine department at the Radiation and Isotopes Center of Khartoum (RICK) during 2003-2005 the aim of the study was explore and define the dimensions of a problem which was regarded as urgent by the people working in the field of nuclear medicine in Sudan. The problem concerned the quality of technitium-labelled radiopharmaceuticals which are used in more than 90% of the nuclear medicine imaging studies performed in nuclear medicine. Impure 9 ''9''m Tc-labelled radiopharmaceuticals may create problems, and could lead to false diagnosis. These agents must be tested for determination of the levels of radionuclides, radiochemical and chemicals, before administration to patients. They should also be sterile and pyrogen-free. A number of data collection methods, were used by the researcher for adequate exploration of the dimensions of the problem including interviews, questionnaires and close observations to all activities related to the preparation of radiopharmaceuticals in the hot laboratory. Information concerning all the aspects of quality assurance were collected. These aspects were management and organisation of the work, equipment and tools, knowledge and practical experience of the staff members and methods of preparation and administration of the radioactive agents. Data from different sources were then compared with observation results for more validation and finally lead to the following results: All the quality control tests were not normally performed in the department, therefore the levels of impurities in these agents were not exactly determined, moreover these preparations were subject to contamination with microorganisms, due to low level of cleanliness at the work area. The study detected a number of defaults which were likely to be the causes behind these problems. These were, bad management and organisation, in availability of equipment, tools and materials necessary for testing

  9. Reliable LC-MS quantitative glycomics using iGlycoMab stable isotope labeled glycans as internal standards.

    Science.gov (United States)

    Zhou, Shiyue; Tello, Nadia; Harvey, Alex; Boyes, Barry; Orlando, Ron; Mechref, Yehia

    2016-06-01

    Glycans have numerous functions in various biological processes and participate in the progress of diseases. Reliable quantitative glycomic profiling techniques could contribute to the understanding of the biological functions of glycans, and lead to the discovery of potential glycan biomarkers for diseases. Although LC-MS is a powerful analytical tool for quantitative glycomics, the variation of ionization efficiency and MS intensity bias are influencing quantitation reliability. Internal standards can be utilized for glycomic quantitation by MS-based methods to reduce variability. In this study, we used stable isotope labeled IgG2b monoclonal antibody, iGlycoMab, as an internal standard to reduce potential for errors and to reduce variabililty due to sample digestion, derivatization, and fluctuation of nanoESI efficiency in the LC-MS analysis of permethylated N-glycans released from model glycoproteins, human blood serum, and breast cancer cell line. We observed an unanticipated degradation of isotope labeled glycans, tracked a source of such degradation, and optimized a sample preparation protocol to minimize degradation of the internal standard glycans. All results indicated the effectiveness of using iGlycoMab to minimize errors originating from sample handling and instruments. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Evaluation of Isotope 32P Method to Mark Culex pipiens (Diptera: Culicidae in a Laboratory

    Directory of Open Access Journals (Sweden)

    Chongxing Zhang

    2016-01-01

    Full Text Available Background: The aim of the current study was to develop a marking technique as an internal marker to mark post blood meal mosquitoes by using stable phosphate isotope 32P and determine the optimal concentration of it.Methods: An isotonic physiological saline solution, containing different concentration of radioactive isotope 32P-labeled disodium phosphate (Na2H32PO4 was injected into rabbits via the jugular vein in the laboratory. Emerged Cx. pipiens were marked after feeding on rabbit. At the same time, the labeled conditions of emerged Cx. pipiens were also measured by placing feces of No. 6 rabbit into containers with mosquito larvae and pupae inside.Results: According to the label condition of Cx. pipiens after taking blood and the effect of different dosage Na2H32PO4 on rabbit health, the optimal concentration of radioactive isotope was determined, that is, 0.1211 mCi/kg. By placing feces of No. 6 rabbit into containers with mosquito larvae and pupae inside, the emerged mosquitoes were also labeled. Therefore, feeding mosquitoes on the animal injected with radioactive Na2H32PO4 was more prac­tical for detecting and tracing mosquitoes.Conclusion: The method was less time-consuming, more sensitive and safer. This marking method will facilitate post-bloodmeal studies of mosquitoes and other blood-sucking insects.

  11. Method of deuterium isotope separation and enrichment

    International Nuclear Information System (INIS)

    Benson, S.W.

    1980-01-01

    A method of deuterium isotope separation and enrichment using infrared laser technology in combination with chemical processes for treating and recycling the unreacted and deuterium-depleted starting materials is described. Organic molecules of the formula RX (where R is an ethyl, isopropyl, t-butyl, or cyclopentenyl group and X is F, Cl, Br or OH) containing a normal abundance of hydrogen and deuterium are exposed to intense laser infrared radiation. An olefin containing deuterium (olefin D) will be formed, along with HX. The enriched olefin D can be stripped from the depleted stream of RX and HX, and can be burned to form enriched water or pyrolyzed to produce hydrogen gas with elevated deuterium content. The depleted RX is decomposed to olefins and RX, catalytically exchanged with normal water to restore the deuterium content to natural levels, and recombined to form RX which can be recycled. (LL)

  12. Differential isotope dansylation labeling combined with liquid chromatography mass spectrometry for quantification of intact and N-terminal truncated proteins

    International Nuclear Information System (INIS)

    Tang, Yanan; Li, Liang

    2013-01-01

    Graphical abstract: -- Highlights: •LC–MS was developed for quantifying protein mixtures containing both intact and N-terminal truncated proteins. • 12 C 2 -Dansylation of the N-terminal amino acid of proteins was done first, followed by microwave-assisted acid hydrolysis. •The released 12 C 2 -dansyl labeled N-terminal amino acid was quantified using 13 C 2 -dansyl labeled amino acid standards. •The method provided accurate and precise results for quantifying intact and N-terminal truncated proteins within 8 h. -- Abstract: The N-terminal amino acids of proteins are important structure units for maintaining the biological function, localization, and interaction networks of proteins. Under different biological conditions, one or several N-terminal amino acids could be cleaved from an intact protein due to processes, such as proteolysis, resulting in the change of protein properties. Thus, the ability to quantify the N-terminal truncated forms of proteins is of great importance, particularly in the area of development and production of protein-based drugs where the relative quantity of the intact protein and its truncated form needs to be monitored. In this work, we describe a rapid method for absolute quantification of protein mixtures containing intact and N-terminal truncated proteins. This method is based on dansylation labeling of the N-terminal amino acids of proteins, followed by microwave-assisted acid hydrolysis of the proteins into amino acids. It is shown that dansyl labeled amino acids are stable in acidic conditions and can be quantified by liquid chromatography mass spectrometry (LC–MS) with the use of isotope analog standards

  13. Differential isotope dansylation labeling combined with liquid chromatography mass spectrometry for quantification of intact and N-terminal truncated proteins

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Yanan; Li, Liang, E-mail: Liang.Li@ualberta.ca

    2013-08-20

    Graphical abstract: -- Highlights: •LC–MS was developed for quantifying protein mixtures containing both intact and N-terminal truncated proteins. •{sup 12}C{sub 2}-Dansylation of the N-terminal amino acid of proteins was done first, followed by microwave-assisted acid hydrolysis. •The released {sup 12}C{sub 2}-dansyl labeled N-terminal amino acid was quantified using {sup 13}C{sub 2}-dansyl labeled amino acid standards. •The method provided accurate and precise results for quantifying intact and N-terminal truncated proteins within 8 h. -- Abstract: The N-terminal amino acids of proteins are important structure units for maintaining the biological function, localization, and interaction networks of proteins. Under different biological conditions, one or several N-terminal amino acids could be cleaved from an intact protein due to processes, such as proteolysis, resulting in the change of protein properties. Thus, the ability to quantify the N-terminal truncated forms of proteins is of great importance, particularly in the area of development and production of protein-based drugs where the relative quantity of the intact protein and its truncated form needs to be monitored. In this work, we describe a rapid method for absolute quantification of protein mixtures containing intact and N-terminal truncated proteins. This method is based on dansylation labeling of the N-terminal amino acids of proteins, followed by microwave-assisted acid hydrolysis of the proteins into amino acids. It is shown that dansyl labeled amino acids are stable in acidic conditions and can be quantified by liquid chromatography mass spectrometry (LC–MS) with the use of isotope analog standards.

  14. Simultaneous determination of glucose turnover, alanine turnover, and gluconeogenesis in human using a double stable-isotope-labeled tracer infusion and gas chromatography-mass spectrometry analysis

    International Nuclear Information System (INIS)

    Martineau, A.; Lecavalier, L.; Falardeau, P.; Chiasson, J.L.

    1985-01-01

    We have developed and validated a new method to measure simultaneously glucose turnover, alanine turnover, and gluconeogenesis in human, in steady and non-steady states, using a double stable-isotope-labeled tracer infusion and GC-MS analysis. The method is based on the concomitant infusion and dilution of D-[2,3,4,6,6-2H5]glucose and L-[1,2,3-13C3]alanine. The choice of the tracers was done on the basis of a minimal overlap between the ions of interest and those arising from natural isotopic abundances. Alanine was chosen as the gluconeogenic substrate because it is the major gluconeogenic amino acid extracted by the liver and, with lactate, constitutes the bulk of the gluconeogenic precursors. The method was validated by comparing the results obtained during simultaneous infusion of trace amounts of both stable isotope labeled compounds with the radioactive tracers (D-[3-3H]glucose and L-[1,2,3-14C3]alanine) in a normal and a diabetic subject; the radiolabeled tracers were used as the accepted reference procedure. A slight overestimation of glucose turnover (7.3 versus 6.8 in normal and 10.8 versus 9.2 mumol/kg min in diabetic subject) was noticed when the stable isotope-labeled tracers were used. For the basal turnover rate of alanine, similar values were obtained with both methods (6.2 mumol/kg min). For gluconeogenesis, higher values were observed in the basal state with the stable isotopes (0.42 versus 0.21 mumol/kg min); however, these differences disappeared in the postprandial period after the ingestion of a mixed meal. Despite those minor differences, the overall correlation with the reference method was excellent for glucose turnover (r = 0.87) and gluconeogenesis (r = 0.86)

  15. ATTA - A new method of ultrasensitive isotope trace analysis

    International Nuclear Information System (INIS)

    Bailey, K.; Chen, C.Y.; Du, X.; Li, Y.M.; Lu, Z.-T.; O'Connor, T.P.; Young, L.

    2000-01-01

    A new method of ultrasensitive isotope trace analysis has been developed. This method, based on the technique of laser manipulation of neutral atoms, has been used to count individual 85 Kr and 81 Kr atoms present in a natural krypton gas sample with isotopic abundances in the range of 10 -11 and 10 -13 , respectively. This method is free of contamination from other isotopes and elements and can be applied to various different isotope tracers for a wide range of applications. The demonstrated detection efficiency is 1x10 -7 . System improvements could increase the efficiency by many orders of magnitude

  16. One-carbon 13C-labeled synthetic intermediates. Comparison and evaluation of preparative methods

    International Nuclear Information System (INIS)

    Ott, D.G.

    1978-01-01

    Frequently the biggest stumbling block to the synthesis of a structurally complex labeled compound is obtaining the required low molecular weight, structurally simple, isotopic intermediates. Selection of a particular scheme from various alternatives depends on the available capabilities and quantity of product desired, as well as on anticipated future requirements and need for related compounds. Many of the newer reagents for organic synthesis can be applied effectively to isotopic preparations with improvements of yields and simplification of procedures compared to established classical methods. New routes developed for higher molecular weight compounds are sometimes not directly adaptable to the one-carbon analogs, either because of isolation difficulties occasioned by physical properties or by chemical reactivities peculiar to their being first members of homologous series. Various routes for preparation of carbon-13 labeled methanol, formaldehyde, and cyanide are compared

  17. A guide through the computational analysis of isotope-labeled mass spectrometry-based quantitative proteomics data: an application study

    Directory of Open Access Journals (Sweden)

    Haußmann Ute

    2011-06-01

    Full Text Available Abstract Background Mass spectrometry-based proteomics has reached a stage where it is possible to comprehensively analyze the whole proteome of a cell in one experiment. Here, the employment of stable isotopes has become a standard technique to yield relative abundance values of proteins. In recent times, more and more experiments are conducted that depict not only a static image of the up- or down-regulated proteins at a distinct time point but instead compare developmental stages of an organism or varying experimental conditions. Results Although the scientific questions behind these experiments are of course manifold, there are, nevertheless, two questions that commonly arise: 1 which proteins are differentially regulated regarding the selected experimental conditions, and 2 are there groups of proteins that show similar abundance ratios, indicating that they have a similar turnover? We give advice on how these two questions can be answered and comprehensively compare a variety of commonly applied computational methods and their outcomes. Conclusions This work provides guidance through the jungle of computational methods to analyze mass spectrometry-based isotope-labeled datasets and recommends an effective and easy-to-use evaluation strategy. We demonstrate our approach with three recently published datasets on Bacillus subtilis 12 and Corynebacterium glutamicum 3. Special focus is placed on the application and validation of cluster analysis methods. All applied methods were implemented within the rich internet application QuPE 4. Results can be found at http://qupe.cebitec.uni-bielefeld.de.

  18. Carbon allocation belowground in Pinus pinaster using stable carbon isotope pulse labeling technique

    Science.gov (United States)

    Dannoura, M.; Bosc, A.; Chipeaux, C.; Sartore, M.; Lambrot, C.; Trichet, P.; Bakker, M.; Loustau, D.; Epron, D.

    2010-12-01

    Carbon allocation belowground competes with aboveground growth and biomass production. In the other hand, it contributes to resource acquisition such as nutrient, water and carbon sequestration in soil. Thus, a better characterization of carbon flow from plant to soil and its residence time within each compartment is an important issue for understanding and modeling forest ecosystem carbon budget. 13C pulse labeling of whole crown was conducted at 4 seasons to study the fate of assimilated carbon by photosynthesis into the root on 12 year old Pinus pinaster planted in the INRA domain of Pierroton. Maritime pine is the most widely planted species in South-West Europe. Stem, root and soil CO2 effluxes and their isotope composition were measured continuously by tunable diode laser absorption spectroscopy with a trace gas analyzer (TGA 100A; Campbell Scientific) coupled to flow-through chambers. 13CO2 recovery and peak were observed in respiration of each compartment after labeling. It appeared sequentially from top of stem to bottom, and to coarse root. The maximum velocity of carbon transfer was calculated as the difference in time lag of recovery between two positions on the trunk or on the root. It ranged between 0.08-0.2 m h-1 in stem and between 0.04-0.12 m h-1 in coarse root. This velocity was higher in warmer season, and the difference between time lag of recovery and peak increased after first frost. Photosynthates arrived underground 1.5 to 5 days after labeling, at similar time in soil CO2 effluxes and coarse root respiration. 0.08-1.4 g of carbon was respired per tree during first 20 days following labeling. It presented 0.6 -10% of 13C used for labeling and it is strongly related to seasons. The isotope signal was detected in fine root organs and microbial biomass by periodical core sampling. The peak was observed 6 days after labeling in early summer while it was delayed more than 10 days in autumn and winter with less amount of carbon allocated

  19. Method and apparatus for separating uranium isotopes

    International Nuclear Information System (INIS)

    Bernstein, E.R.

    1977-01-01

    A uranium compound in the solid phase (uranium borohydride four) is subjected to radiation of a first predetermined frequency that excites the uranium-235 isotope-bearing molecules but not the uranium-238 isotope-bearing molecules. The compound is simultaneously subjected to radiation of a second predetermined frequency which causes the excited uranium-235 isotope-bearing molecules to chemically decompose but which does not affect the uranium-238 isotope-bearing molecules. Sufficient heat is then applied to the irradiated compound in the solid phase to vaporize the non-decomposed uranium-238 isotope-bearing molecules but not the decomposed uranium-235 isotope-bearing molecules, thereby physically separating the uranium-235 isotope-bearing molecules from the uranium-238 isotope-bearing molecules. The uranium compound sample in the solid phase is deposited or grown in an elongated tube supported within a dewar vessel having a clear optical path tail section surrounded by a coolant. Two sources of radiation are focused on the uranium compound sample. A heating element is attached to the elongated tube to vaporize the irradiated compound

  20. Gas isotope separation method using plasma sheet

    International Nuclear Information System (INIS)

    Takayama, K.; Takagi, K.; Fukvi, R.

    1988-03-01

    A high frequency electric field is applied to a plasma sheet with a frequency equal to the cyclotronic frequency of the ions to be separated. Because of resonance the cyclotronic radius of the isotope has increased and the electric charge is eliminated by collision with a separator and the isotope is separated in neutral particles [fr

  1. Method of isotope separation by chemi-ionization

    International Nuclear Information System (INIS)

    Wexler, S.; Young, C.E.

    1977-01-01

    A method is disclosed for separating isotopes in an isotopic mixture by aerodynamically accelerating a gaseous compound to form a jet of molecules, and passing the jet through a stream of electron donor atoms whereby an electron transfer takes place, thus forming negative ions of the molecules. The molecular ions are then passed through a radiofrequency quadrupole mass filter to separate the specific isotopes. cThis method may be used for any compounds having a sufficiently high electron affinity to permit negative ion formation, and is especially useful for the separation of plutonium and uranium isotopes. 10 claims, 1 figure

  2. Basic methods of isotope analysis; Osnovnye metody analiza izotopov

    Energy Technology Data Exchange (ETDEWEB)

    Ochkin, A V; Rozenkevich, M B

    2000-07-01

    The bases of the most applied methods of the isotope analysis are briefly presented. The possibilities and analytical characteristics of the mass-spectrometric, spectral, radiochemical and special methods of the isotope analysis, including application of the magnetic resonance, chromatography and refractometry, are considered.

  3. Method and apparatus for isotope separation from a gas stream

    International Nuclear Information System (INIS)

    Szoke, A.

    1978-01-01

    A method and apparatus are described for isotope separation and in particular for separating the desired isotope from the gas in which it is contained by irradiating it with a laser. The laser selectively provides kinetic energy to the isotope through inelastic events, monomolecular or bimolecular, in order to cause it to segregate within or fly out of the gas stream in which it is contained

  4. The evaluation of new and isotopically labeled isoindoline nitroxides and an azaphenalene nitroxide for EPR oximetry

    Science.gov (United States)

    Khan, Nadeem; Blinco, James P.; Bottle, Steven E.; Hosokawa, Kazuyuki; Swartz, Harold M.; Micallef, Aaron S.

    2011-01-01

    Isoindoline nitroxides are potentially useful probes for viable biological systems, exhibiting low cytotoxicity, moderate rates of biological reduction and favorable Electron Paramagnetic Resonance (EPR) characteristics. We have evaluated the anionic (5-carboxy-1,1,3,3-tetramethylisoindolin-2-yloxyl; CTMIO), cationic (5-(N,N,N-trimethylammonio)-1,1,3,3-tetramethylisoindolin-2-yloxyl iodide, QATMIO) and neutral (1,1,3,3-tetramethylisoindolin-2-yloxyl; TMIO) nitroxides and their isotopically labeled analogues (2H12- and/or 2H12-15N-labeled) as potential EPR oximetry probes. An active ester analogue of CTMIO, designed to localize intracellularly, and the azaphenalene nitroxide 1,1,3,3-tetramethyl-2,3-dihydro-2-azaphenalen-2-yloxyl (TMAO) were also studied. While the EPR spectra of the unlabeled nitroxides exhibit high sensitivity to O2 concentration, deuteration resulted in a loss of superhyperfine features and a subsequent reduction in O2 sensitivity. Labeling the nitroxides with 15N increased the signal intensity and this may be useful in decreasing the detection limits for in vivo measurements. The active ester nitroxide showed approximately 6% intracellular localization and low cytotoxicity. The EPR spectra of TMAO nitroxide indicated an increased rigidity in the nitroxide ring, due to dibenzo-annulation. PMID:21665499

  5. Labeling pharmaceuticals with radioactive isotopes. Technical progress report, December 1, 1975--November 30, 1976

    International Nuclear Information System (INIS)

    Blau, M.; Bender, M.A.

    1976-01-01

    The purpose of this research is to prepare iodo- and bromo-aliphatic amino acid analogs labeled with γ-emitting isotopes ( 131 I, 123 I and 77 Br) for possible use as pancreas localizing agents. Studies on the halogen exchange reaction (I- for Cl-) for the synthesis of β-iodo-α-aminobutyric acid (a valine analog) have suggested that the iodo compound was formed initially. However, the desired compound cannot be isolated because of its chemical instability. Distribution studies in rats with the crude halogen exchange reaction mixture confirmed this finding. Studies on the addition of hydrogen iodine to allylglycine under various conditions for the synthesis of γ-iodo-α-aminopentanoic acid (a leucine analog) suffered the same obstacle; the chemical instability of the desired iodo compound precludes isolation and characterization. Convinced that the iodo analogs were too unstable for use as practical localizing agents, we turned to the possible use of Br for CH 3 substituted amino acids. The 14 C labeled β-bromo-α-aminobutyric acid methyl ester was synthesized. This methyl ester will be hydrolyzed and the distribution of free amino acid will be studied. Labeled with 77 Br this compound might be useful for pancreas localization

  6. Method and apparatus for separating isotopes

    International Nuclear Information System (INIS)

    Harris, S.E.

    1976-01-01

    Isotope separation is achieved between species A and B having an absorption resonance separated by an isotopic shift by selectively exciting a portion of species A using a tunable photon source of narrow emission line with and subsequently causing collisions with an optically excited third species to selectively ionize the excited portion of species A. When ionized, species A is easily separated by any technique, using its ionized condition to distinguish it from species B. 18 claims, 3 drawing figures

  7. Technetium-99m labeled radiodiagnostic agents and method of preparation

    International Nuclear Information System (INIS)

    1976-01-01

    A method of preparing improved technetium-99m labelled radiodiagnostic agents by reducing sup(99m)Tc-pertechnetate with stannous tartrate is given. Human serum albumine (HSA) and 1-hydroxyethylidene-1,1-disodiumphosphonate (HEDSPA), which are useful in scintigraphic examinations of the lung and bone, were labelled in this way

  8. Affordable uniform isotope labeling with {sup 2}H, {sup 13}C and {sup 15}N in insect cells

    Energy Technology Data Exchange (ETDEWEB)

    Sitarska, Agnieszka; Skora, Lukasz; Klopp, Julia; Roest, Susan; Fernández, César; Shrestha, Binesh; Gossert, Alvar D., E-mail: alvar.gossert@novartis.com [Novartis Institutes for BioMedical Research (Switzerland)

    2015-06-15

    For a wide range of proteins of high interest, the major obstacle for NMR studies is the lack of an affordable eukaryotic expression system for isotope labeling. Here, a simple and affordable protocol is presented to produce uniform labeled proteins in the most prevalent eukaryotic expression system for structural biology, namely Spodoptera frugiperda insect cells. Incorporation levels of 80 % can be achieved for {sup 15}N and {sup 13}C with yields comparable to expression in full media. For {sup 2}H,{sup 15}N and {sup 2}H,{sup 13}C,{sup 15}N labeling, incorporation is only slightly lower with 75 and 73 %, respectively, and yields are typically twofold reduced. The media were optimized for isotope incorporation, reproducibility, simplicity and cost. High isotope incorporation levels for all labeling patterns are achieved by using labeled algal amino acid extracts and exploiting well-known biochemical pathways. The final formulation consists of just five commercially available components, at costs 12-fold lower than labeling media from vendors. The approach was applied to several cytosolic and secreted target proteins.

  9. Stem cell monitoring with a direct or indirect labeling method

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Min Hwan; Lee, Yong Jin [Molecular Imaging Research Center, Korea Institute of Radiological and Medical Sciences (KIRAMS), Seoul (Korea, Republic of)

    2016-12-15

    The molecular imaging techniques allow monitoring of the transplanted cells in the same individuals over time, from early localization to the survival, migration, and differentiation. Generally, there are two methods of stem cell labeling: direct and indirect labeling methods. The direct labeling method introduces a labeling agent into the cell, which is stably incorporated or attached to the cells prior to transplantation. Direct labeling of cells with radionuclides is a simple method with relatively fewer adverse events related to genetic responses. However, it can only allow short-term distribution of transplanted cells because of the decreasing imaging signal with radiodecay, according to the physical half-lives, or the signal becomes more diffuse with cell division and dispersion. The indirect labeling method is based on the expression of a reporter gene transduced into the cell before transplantation, which is then visualized upon the injection of an appropriate probe or substrate. In this review, various imaging strategies to monitor the survival and behavior change of transplanted stem cells are covered. Taking these new approaches together, the direct and indirect labeling methods may provide new insights on the roles of in vivo stem cell monitoring, from bench to bedside.

  10. Method and device for the enrichment of isotopes

    International Nuclear Information System (INIS)

    Stehle, H.

    1976-01-01

    A variation of a method for isotope enrichment by laser radiation is proposed which improves the selectivity and with it the economy of the method by eliminating undesired reactions caused by thermal activation. The method according to the invention is applied discontinuously in three steps: The isotope mixture and the reacting agents are fed to a vessel, a laser beam is passed through, and the contents are emptied into a vacuum tank while expanding adiabatically. The time steps are controlled. The method is explained using the example of separating an isotope mixture of UF 6 . (UWI) [de

  11. Adaptation of a Commonly Used, Chemically Defined Medium for Human Embryonic Stem Cells to Stable Isotope Labeling with Amino Acids in Cell Culture

    DEFF Research Database (Denmark)

    Liberski, A. R.; Al-Noubi, M. N.; Rahman, Z. H.

    2013-01-01

    Metabolic labeling with stable isotopes is a prominent technique for comparative quantitative proteomics, and stable isotope labeling with amino acids in cell culture (SILAC) is the most commonly used approach. SILAC is, however, traditionally limited to simple tissue culture regimens and only ra...

  12. Exploration of new tritium labelling methods

    International Nuclear Information System (INIS)

    Andres, H.; Jaiswal, D.K.; Morimoto, H.; Saljoughian, M.; Than, C.; Willimas, P.G.; Zippi, E.M.

    1997-01-01

    Full text: A great deal of elegant chemistry is available for hydride transfer reactions, and could be adapted for tritium labelling. Nevertheless, most high level tritiation reactions still involve either hydrogenation (alkene or alkyne precursor) or catalytic dehalogenation. In the last decade we have endeavored to propose and popularize alternative labelling techniques and reagents, including: i) the synthesis of new precursors for the production of methyl iodide; ii) the synthesis of methylene diiodide; iii) the production and use of T 2 O, and solvents made from it, eg. CH 3 COOT, CF 3 COOT; iv) high specific activity hydride reagents, eg. LiAIT 4 , LiEt 3 BT; (Bu n ) 3 SnT, ZrCp 2 CIT, LiT, Li(OCH 3 ) 3 BT, Ph 2 SiT 2 , BT 3 -THF, Li/Na/KBT 4 ; v) reduction with diimide; vi) use of T 2 O in special reactions such as the Shapiro reaction and Brook rearrangement; and vii) developments of a new acetylation reagent. We have also initiated and continued a number of innovative applications of tritium NMR spectroscopy. Many of these projects have grown out of User or Collaborator requirements at the NTLF. We regard this stimulus to develop and refine both tritiation and NMR techniques as healthy and challenging

  13. Isotope labeling-based quantitative proteomics of developing seeds of castor oil seed (Ricinus communis L.)

    DEFF Research Database (Denmark)

    Nogueira, Fábio C S; Palmisano, Giuseppe; Schwämmle, Veit

    2013-01-01

    In this study, we used a mass spectrometry-based quantification approach employing isotopic (ICPL) and isobaric (iTRAQ) labeling to investigate the pattern of protein deposition during castor oil seed (Ricinus communis L.) development, including that of proteins involved in fatty acid metabolism...... give important insights into certain aspects of the biology of castor oil seed development such as carbon flow, anabolism, and catabolism of fatty acid and the pattern of deposition of SSPs, toxins, and allergens such as ricin and 2S albumins. We also found, for the first time, some genes of SSP......, seed-storage proteins (SSPs), toxins, and allergens. Additionally, we have used off-line hydrophilic interaction chromatography (HILIC) as a step of peptide fractionation preceding the reverse-phase nanoLC coupled to a LTQ Orbitrap. We were able to identify a total of 1875 proteins, and from these 1748...

  14. Bacterial expression and one-step purification of an isotope-labeled heterotrimeric G-protein {alpha}-subunit

    Energy Technology Data Exchange (ETDEWEB)

    Abdulaev, Najmoutin G. [University of Maryland Biotechnology Institute, Center for Advanced Research in Biotechnology (United States); Zhang Cheng; Dinh, Andy [University of Texas Health Science Center, Center for Membrane Biology, Department of Biochemistry and Molecular Biology (United States); Ngo, Tony; Bryan, Philip N. [University of Maryland Biotechnology Institute, Center for Advanced Research in Biotechnology (United States); Brabazon, Danielle M. [Loyola College in Maryland, Department of Chemistry (United States); Marino, John P. [University of Maryland Biotechnology Institute, Center for Advanced Research in Biotechnology (United States)], E-mail: marino@carb.nist.gov; Ridge, Kevin D. [University of Texas Health Science Center, Center for Membrane Biology, Department of Biochemistry and Molecular Biology (United States)

    2005-05-15

    Heterologous expression systems are often employed to generate sufficient quantities of isotope-labeled proteins for high-resolution NMR studies. Recently, the interaction between the prodomain region of subtilisin and an active, mutant form of the mature enzyme has been exploited to develop a cleavable affinity tag fusion system for one-step generation and purification of full-length soluble proteins obtained by inducible prokaryotic expression. As a first step towards applying high-resolution NMR methods to study heterotrimeric G-protein {alpha}-subunit (G{sub {alpha}}) conformation and dynamics, the utility of this subtilisin prodomain fusion system for expressing and purifying an isotope-labeled G{sub {alpha}} chimera ({approx}40 kDa polypeptide) has been tested. The results show that a prodomain fused G{sub {alpha}} chimera can be expressed to levels approaching 6-8 mg/l in minimal media and that the processed, mature protein exhibits properties similar to those of G{sub {alpha}} isolated from natural sources. To assay for the functional integrity of the purified G{sub {alpha}} chimera at NMR concentrations and probe for changes in the structure and dynamics of G{sub {alpha}} that result from activation, {sup 15}N-HSQC spectra of the GDP/Mg{sup 2+} bound form of G{sub {alpha}} obtained in the absence and presence of aluminum fluoride, a well known activator of the GDP bound state, have been acquired. Comparisons of the {sup 15}N-HSQC spectra reveals a number of changes in chemical shifts of the {sup 1}HN, {sup 15}N crosspeaks that are discussed with respect to expected changes in the protein conformation associated with G{sub {alpha}} activation.

  15. Bacterial expression and one-step purification of an isotope-labeled heterotrimeric G-protein α-subunit

    International Nuclear Information System (INIS)

    Abdulaev, Najmoutin G.; Zhang Cheng; Dinh, Andy; Ngo, Tony; Bryan, Philip N.; Brabazon, Danielle M.; Marino, John P.; Ridge, Kevin D.

    2005-01-01

    Heterologous expression systems are often employed to generate sufficient quantities of isotope-labeled proteins for high-resolution NMR studies. Recently, the interaction between the prodomain region of subtilisin and an active, mutant form of the mature enzyme has been exploited to develop a cleavable affinity tag fusion system for one-step generation and purification of full-length soluble proteins obtained by inducible prokaryotic expression. As a first step towards applying high-resolution NMR methods to study heterotrimeric G-protein α-subunit (G α ) conformation and dynamics, the utility of this subtilisin prodomain fusion system for expressing and purifying an isotope-labeled G α chimera (∼40 kDa polypeptide) has been tested. The results show that a prodomain fused G α chimera can be expressed to levels approaching 6-8 mg/l in minimal media and that the processed, mature protein exhibits properties similar to those of G α isolated from natural sources. To assay for the functional integrity of the purified G α chimera at NMR concentrations and probe for changes in the structure and dynamics of G α that result from activation, 15 N-HSQC spectra of the GDP/Mg 2+ bound form of G α obtained in the absence and presence of aluminum fluoride, a well known activator of the GDP bound state, have been acquired. Comparisons of the 15 N-HSQC spectra reveals a number of changes in chemical shifts of the 1 HN, 15 N crosspeaks that are discussed with respect to expected changes in the protein conformation associated with G α activation

  16. Technetium-99m labeled radiodiagnostic agents and method of preparation

    International Nuclear Information System (INIS)

    Molinski, V.J.; Wilczewski, J.A.

    1977-01-01

    A method of preparing improved technetium-99m labeled radiodiagnostic agents by reducing technetium-99m with stannous tartrate is described. Such radiodiagnostic agents are useful in scintigraphic examinations of the bone and lung

  17. Comments on “innovative method for carbon dioxide determination in human postmortem cardiac gas samples using headspace-gas chromatography–mass spectrometry and stable labeled isotope as internal standard” by Varlet et al

    Energy Technology Data Exchange (ETDEWEB)

    Saffaj, T., E-mail: taoufiq.saffaj@usmba.ac.ma [Centre Universitaire Régional d’Interface, Université Sidi Mohamed Ben Abdellah, BP 2626, route d’Imouzzer, Fes Morocco (Morocco); Laboratoire de Chimie Organique Appliquée, Faculté des Sciences et Techniques, BP 2202, route d’Imouzzer, Fes Morocco (Morocco); Ihssane, B. [Laboratoire de Chimie Organique Appliquée, Faculté des Sciences et Techniques, BP 2202, route d’Imouzzer, Fes Morocco (Morocco)

    2014-01-31

    Graphical abstract: -- Highlights: •We comment on the validation results of the method for CO{sub 2} analysis in cardiac samples. •We demonstrate that the total error of the method exceeds the acceptance limits. •We show that the method is not valid according to the new SFSTP guideline. -- Abstract: Varlet et al. recently proposed a headspace-gas chromatography–mass spectrometry (HS-GC–MS) method applicable for the routine determination of CO{sub 2} in gaseous biologic matrices. This developed bioanalytical method was fully validated according to the SFSTP 1997 guidelines using the accuracy profile as a graphical decision-making tool.In this letter, we discuss the validity of HS-GC–MS method based on the newest SFSTP guideline. In particular, we demonstrate by the estimation of the β-expectation tolerance interval that the error total exceeds the acceptance limits (30%) for the second concentration level (0.5 μmol mL{sup −1} vial HS). Furthermore, we show through the risk profile that the probability to have future results inside the ±30% acceptance limits is smaller than 95%.

  18. Intrinsic mineral labeling of edible plants: methods and uses

    International Nuclear Information System (INIS)

    Weaver, C.M.

    1985-01-01

    The fate of minerals can be conveniently studied through intrinsic labeling techniques. The mineral of interest is biologically incorporated into the food in a form that can be distinguished analytically from the natural form of the element. Radiolabels have traditionally been used to study such problems as the uptake of minerals by plants, the gross and subcellular mineral distribution in plant tissues, the form and associations of the deposited mineral, and the bioavailability of minerals to animals and humans. The use of stable (nonradioactive) isotopes as a label offers the potential of safely studying bioavailability of minerals from individual foods in human population groups of all ages using foods processed in normal food handling and processing facilities. 114 references

  19. Closing the gap between T-cell life span estimates from stable isotope-labeling studies in mice and humans

    NARCIS (Netherlands)

    Westera, Liset; Drylewicz, Julia; den Braber, Ineke; Mugwagwa, Tendai; van der Maas, Iris; Kwast, Lydia; Volman, Thomas; van de Weg-Schrijver, Elise H. R.; Bartha, István; Spierenburg, Gerrit; Gaiser, Koos; Ackermans, Mariëtte T.; Asquith, Becca; de Boer, Rob J.; Tesselaar, Kiki; Borghans, José A. M.

    2013-01-01

    Quantitative knowledge of the turnover of different leukocyte populations is a key to our understanding of immune function in health and disease. Much progress has been made thanks to the introduction of stable isotope labeling, the state-of-the-art technique for in vivo quantification of cellular

  20. Isotope labelling study of CO oxidation-assisted epoxidation of propene. Implications for oxygen activation on Au catalysts.

    Science.gov (United States)

    Jiang, Jian; Oxford, Sean M; Fu, Baosong; Kung, Mayfair C; Kung, Harold H; Ma, Jiantai

    2010-06-07

    (18)O isotope labelling studies of the CO oxidation-assisted epoxidation of propene, catalyzed by a mixture of Au/TiO(2) and TS-1, using a methanol-H(2)O solvent showed the O in the epoxide was exclusively from O(2) and not H(2)O or methanol.

  1. Isotope exchange reactions of the hydrogen H-5 of selected pyrimidine derivatives and the preparation of tritium-labeled pyrimidines

    Czech Academy of Sciences Publication Activity Database

    Dračínský, Martin; Jansa, Petr; Elbert, Tomáš

    2011-01-01

    Roč. 76, č. 12 (2011), s. 1567-1577 ISSN 0010-0765 R&D Projects: GA AV ČR KJB400550903; GA MŠk 1M0508 Institutional research plan: CEZ:AV0Z40550506 Keywords : isotopic labeling * NMR spectroscopy * nucleobases * pyrimidines Subject RIV: CC - Organic Chemistry Impact factor: 1.283, year: 2011

  2. Cross-Course Collaboration in the Undergraduate Chemistry Curriculum: Isotopic Labeling with Sodium Borodeuteride in the Introductory Organic Chemistry Laboratory

    Science.gov (United States)

    Kjonaas, Richard A.; Fitch, Richard W.; Noll, Robert J.

    2017-01-01

    A microscale isotopic labeling experiment is described for the introductory organic chemistry laboratory course wherein half of the students use sodium borohydride (NaBH[subscript 4]) and the other half use sodium borodeuteride (NaBD[subscript 4]) to reduce acetophenone to 1-phenylethanol and then compare spectral data. The cost is reasonable, and…

  3. Solid state NMR of isotope labelled murine fur: a powerful tool to study atomic level keratin structure and treatment effects

    Energy Technology Data Exchange (ETDEWEB)

    Wong, Wai Ching Veronica; Narkevicius, Aurimas; Chow, Wing Ying; Reid, David G.; Rajan, Rakesh [University of Cambridge, Department of Chemistry (United Kingdom); Brooks, Roger A. [University of Cambridge, Department of Trauma and Orthopaedic Surgery, Addenbrooke’s Hospital (United Kingdom); Green, Maggie [University of Cambridge, Central Biomedical Resources, School of Clinical Medicine (United Kingdom); Duer, Melinda J., E-mail: mjd13@cam.ac.uk [University of Cambridge, Department of Chemistry (United Kingdom)

    2016-10-15

    We have prepared mouse fur extensively {sup 13}C,{sup 15}N-labelled in all amino acid types enabling application of 2D solid state NMR techniques which establish covalent and spatial proximities within, and in favorable cases between, residues. {sup 13}C double quantum–single quantum correlation and proton driven spin diffusion techniques are particularly useful for resolving certain amino acid types. Unlike 1D experiments on isotopically normal material, the 2D methods allow the chemical shifts of entire spin systems of numerous residue types to be determined, particularly those with one or more distinctively shifted atoms such as Gly, Ser, Thr, Tyr, Phe, Val, Leu, Ile and Pro. Also the partial resolution of the amide signals into two signal envelopes comprising of α-helical, and β-sheet/random coil components, enables resolution of otherwise overlapped α-carbon signals into two distinct cross peak families corresponding to these respective secondary structural regions. The increase in resolution conferred by extensive labelling offers new opportunities to study the chemical fate and structural environments of specific atom and amino acid types under the influence of commercial processes, and therapeutic or cosmetic treatments.

  4. Discriminating background from anthropogenic lead by isotopic methods

    International Nuclear Information System (INIS)

    Nelson, B.K.; O'Brien, H.E.

    1995-01-01

    The goal of this pilot project was to evaluate the practicality of using natural variations in the isotopic composition of lead to test for the presence of anthropogenic lead in soil, surface water and ground water. Complex chemical reactions in the environment may cause measured lead concentrations to be ambiguous indicators of anthropogenic lead component. The lead isotope tracer technique has the potential to identify both the presence and proportion of anthropogenic lead in the environment. The tested the lead isotope technique at Eielson Air Force Base, Alaska, on sources of suspected fuel contamination. Although the results are specific to this base, the general technique of using lead isotopes to trace the movement of anthropogenic lead is applicable to other CERCLA sites. The study had four objectives: (1) characterize the natural lead isotope composition of bedrock, stream sediment and soils; (2) characterize the isotopic composition of the contaminant lead derived from fuel; (3) evaluate the sensitivity of the isotopic method to distinguishing between anthropogenic and natural lead in soil and water samples and (4) evaluate the analytical feasibility and accuracy of the method at the Isotope Geochemistry Laboratory at the University of Washington

  5. Determination of the isotope distribution in 14C-labelled hydrocarbons by thermal fragmentation

    International Nuclear Information System (INIS)

    Kopinke, F.D.; Dermietzel, J.; Jockisch, W.; Raeuber, G.

    1986-01-01

    The gas chromatographic analysis of pyrolysis products of properly labelled hydrocarbons allows a definite and quantitative determination of the 14 C-distribution in those compounds. For this purpose a simple, fast, and versatilely applicable method has been developed and described

  6. Theoretical Proof and Empirical Confirmation of a Continuous Labeling Method Using Naturally 13C-Depleted Carbon Dioxide

    Institute of Scientific and Technical Information of China (English)

    Weixin Cheng; Feike A. Dijkstra

    2007-01-01

    Continuous isotope labeling and tracing is often needed to study the transformation, movement, and allocation of carbon in plant-soil systems. However, existing labeling methods have numerous limitations. The present study introduces a new continuous labeling method using naturally 13C-depleted CO2. We theoretically proved that a stable level of 13C-CO2 abundance In a labeling chamber can be maintained by controlling the rate of CO2-free air injection and the rate of ambient airflow with coupling of automatic control of CO2 concentration using a CO2 analyzer. The theoretical results were tested and confirmed in a 54 day experiment in a plant growth chamber. This new continuous labeling method avoids the use of radioactive 14C or expensive 13C-enriched CO2 required by existing methods and therefore eliminates issues of radiation safety or unaffordable isotope cost, as well as creating new opportunities for short- or long-term labeling experiments under a controlled environment.

  7. A diagram retrieval method with multi-label learning

    Science.gov (United States)

    Fu, Songping; Lu, Xiaoqing; Liu, Lu; Qu, Jingwei; Tang, Zhi

    2015-01-01

    In recent years, the retrieval of plane geometry figures (PGFs) has attracted increasing attention in the fields of mathematics education and computer science. However, the high cost of matching complex PGF features leads to the low efficiency of most retrieval systems. This paper proposes an indirect classification method based on multi-label learning, which improves retrieval efficiency by reducing the scope of compare operation from the whole database to small candidate groups. Label correlations among PGFs are taken into account for the multi-label classification task. The primitive feature selection for multi-label learning and the feature description of visual geometric elements are conducted individually to match similar PGFs. The experiment results show the competitive performance of the proposed method compared with existing PGF retrieval methods in terms of both time consumption and retrieval quality.

  8. Development of new and improved labelling procedures for introducing isotopic hydrogen and carbon-11 into organic compounds

    International Nuclear Information System (INIS)

    Al-Qahtani, M.H.S.

    1999-10-01

    New and improved methods for introducing radioisotopic hydrogen (tritium) and carbon (positron-emitting short-lived carbon-11, t 1/ 2 = 20.4 min) into organic molecules for application in biological research have been explored. In Chapter 1 the applications of radioactive isotopes in biological and clinical research is surveyed, with particular emphasis on the value of β-emitting tritium and positron-emitting carbon-11. In Chapter 2 we report the use of the non-radioactive hydrogen isotope, deuterium, as a surrogate for tritium in the development of microwave-enhanced labelling procedures, based on catalytic hydrogen transfer to olefins (e.g. styrene, styrene derivatives, cinnamic acid and its derivatives). Hydrogen or deuterium donors (e.g. formate salts) were used alone or in combination with other sources (e.g. D 2 O). The method was found to give fully hydrogenated products using very short microwave irradiation times (∼ 2 min) and was highly reproducible. Importantly, the method is environmentally clean, as when extended to tritiated formates little or no radioactive waste is produced. In Chapter 3 we explored the labelling of CGP 62349 {3-[1-(R)-[3-(4-methoxybenzyl)phosphinyl-2-(S)-hydroxy-propyl- amino]ethyl]benzoic acid}, a γ-aminobutyric acid type B (GABA B ) receptor antagonist, with carbon-11 in order to provide a prospective radioligand for medical imaging with positron emission tomography (PET). Labelling agents, [ 11 C]iodomethane and [ 11 C]methyl triflate, prepared by improved methods, were used in the rapid methylation of desmethyl-CGP 62349. Substantially higher radiochemical yields (78%) of [ 11 C]CGP 62349 were achieved by the new methods compared to that produced in a previously published procedure (9%). In addition, the use of [ 11 C]methyl triflate rather than [ 11 C]iodomethane has the advantage of giving a high radiochemical yield and a lower amount of carrier. In Chapter 4 we report on the use of [ 11 C]carbon monoxide as a labelling

  9. Radiation oxidation of polypropylene: A solid-state 13C NMR study using selective isotopic labeling

    International Nuclear Information System (INIS)

    Mowery, Daniel M.; Assink, Roger A.; Derzon, Dora K.; Klamo, Sara B.; Bernstein, Robert; Clough, Roger L.

    2007-01-01

    Polypropylene samples, in which the three different carbon atoms along the chain were selectively labeled with carbon-13, were subjected to radiation under inert and air atmospheres, and to post-irradiation exposure in air at various temperatures. By using solid-state 13 C NMR measurements at room temperature, we have been able to identify and quantify the oxidation products. The isotopic labeling provides insight into chemical reaction mechanisms, since oxidation products can be traced back to their positions of origin on the macromolecule. The major products include peroxides and alcohols, both formed at tertiary carbon sites along the chain. Other products include methyl ketones, acids, esters, peresters, and hemiketals formed from reaction at the tertiary carbon, together with in-chain ketones and esters from reaction at the secondary chain carbon. No evidence is found of products arising from reactions at the methyl side chain. Significant temperature-dependent differences are apparent; for example much higher yields of chain-end methyl ketones, which are the indicator product of chain scission, are generated for both elevated temperature irradiation and for post-irradiation treatment at elevated temperatures. Time-dependent plots of yields of the various oxidation products have been obtained under a wide range of conditions, including the post-irradiation oxidation of a sample at room temperature in air that has been monitored for 2 years. Radiation-oxidation products of polypropylene are contrasted to products measured for 13 C-labeled polyethylene in an earlier investigation: the peroxides formed in irradiated polypropylene are remarkably longer lived, the non-peroxidic products are significantly different, and the overall ratios of oxidation products in polypropylene change relatively little as a function of the extent of oxidation

  10. Identification of degradation routes of metamitron in soil microcosms using 13C-isotope labeling.

    Science.gov (United States)

    Wang, Shizong; Miltner, Anja; Nowak, Karolina M

    2017-01-01

    Metamitron is one of the most commonly used herbicide in sugar beet and flower bulb cultures. Numerous laboratory and field studies on sorption and degradation of metamitron were performed. Detailed biodegradation studies in soil using 13 C-isotope labeling are still missing. Therefore, we aimed at providing a detailed turnover mass balance of 13 C 6 -metamitron in soil microcosms over 80 days. In the biotic system, metamitron mineralized rapidly, and 13 CO 2 finally constituted 60% of the initial 13 C 6 -metamitron equivalents. In abiotic control experiments CO 2 rose to only 7.4% of the initial 13 C 6 -metamitron equivalents. The 13 C label from 13 C 6 -metamitron was incorporated into microbial amino acids that were ultimately stabilized in the soil organic matter forming presumably harmless biogenic residues. Finally, 13 C label from 13 C 6 -metamitron was distributed between the 13 CO 2 and the 13 C-biogenic residues indicating nearly complete biodegradation. The parallel increase of 13 C-alanine, 13 C-glutamate and 13 CO 2 indicates that metamitron was initially biodegraded via the desamino-metamitron route suggesting its relevance in the growth metabolism. In later phases of biodegradation, the "Rhodococcus route" was indicated by the low 13 CO 2 evolution and the high relevance of the pyruvate pathway, which aims at biomolecule synthesis and seems to be related to starvation. This is a first report on the detailed degradation route of metamitron in soil. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. Methods for removing radioactive isotopes from contaminated streams

    International Nuclear Information System (INIS)

    Hoy, D.R.; Hickey, T.N.; Spulgis, I.S.; Parish, H.C.

    1979-01-01

    Methods for removing radioactive isotopes from contaminated gas streams for use in atmospheric containment and cleanup systems in nuclear power plants are provided. The methods provide for removal of radioactive isotopes from a first portion of the contaminated stream, separated from the remaining portion of the stream, so that adsorbent used to purify the first portion of the contaminated stream by adsorption of the radioactive isotopes therefrom can be tested to determine the adsorbing efficacy of the generally larger portion of adsorbent used to purify the remaining portion of the stream

  12. New Isotope Analysis Method: Atom Trap Mass Spectrometry

    International Nuclear Information System (INIS)

    Ko, Kwang Hoon; Park, Hyun Min; Han, Jae Min; Kim, Taek Soo; Cha, Yong Ho; Lim, Gwon; Jeong, Do Young

    2011-01-01

    Trace isotope analysis has been an important role in science, archaeological dating, geology, biology and nuclear industry. Some fission products such as Sr-90, Cs-135 and Kr-85 can be released to the environment when nuclear accident occurs and the reprocessing factory operates. Thus, the analysis of artificially produced radioactive isotopes has been of interest in nuclear industry. But it is difficult to detect them due to low natural abundance less then 10 -10 . In general, radio-chemical method has been applied to detect ultra-trace radio isotopes. But this method has disadvantages of long measurement time for long lived radioisotopes and toxic chemical process for the purification. The Accelerator Mass Spectrometer has high isotope selectivity, but the system is huge and its selectivity is affected by isobars. The laser based method, such as RIMS (Resonance Ionization Mass Spectrometry) has the advantage of isobar-effect free characteristics. But the system size is still huge for high isotope selective system. Recently, ATTA (Atom Trap Trace Analysis) has been successfully applied to detect ultra-trace isotope, Kr-81 and Kr-85. ATTA is the isobar-effect free detection with high isotope selectivity and the system size is small. However, it requires steady atomic beam source during detection, and is not allowed simultaneous detection of several isotopes. In this presentation, we introduce new isotope detection method which is a coupled method of Atom Trap Mass Spectrometry (ATMS). We expect that it can overcome the disadvantage of ATTA while it has both advantages of ATTA and mass spectrometer. The basic concept and the system design will be presented. In addition, the experimental status of ATMS will also be presented

  13. Measurement of apolipoprotein E and amyloid β clearance rates in the mouse brain using bolus stable isotope labeling

    Science.gov (United States)

    2012-01-01

    Background Abnormal proteostasis due to alterations in protein turnover has been postulated to play a central role in several neurodegenerative diseases. Therefore, the development of techniques to quantify protein turnover in the brain is critical for understanding the pathogenic mechanisms of these diseases. We have developed a bolus stable isotope-labeling kinetics (SILK) technique coupled with multiple reaction monitoring mass spectrometry to measure the clearance of proteins in the mouse brain. Results Cohorts of mice were pulse labeled with 13 C6-leucine and the brains were isolated after pre-determined time points. The extent of label incorporation was measured over time using mass spectrometry to measure the ratio of labeled to unlabeled apolipoprotein E (apoE) and amyloid β (Aβ). The fractional clearance rate (FCR) was then calculated by analyzing the time course of disappearance for the labeled protein species. To validate the technique, apoE clearance was measured in mice that overexpress the low-density lipoprotein receptor (LDLR). The FCR in these mice was 2.7-fold faster than wild-type mice. To demonstrate the potential of this technique for understanding the pathogenesis of neurodegenerative disease, we applied our SILK technique to determine the effect of ATP binding cassette A1 (ABCA1) on both apoE and Aβ clearance. ABCA1 had previously been shown to regulate both the amount of apoE in the brain, along with the extent of Aβ deposition, and represents a potential molecular target for lowering brain amyloid levels in Alzheimer's disease patients. The FCR of apoE was increased by 1.9- and 1.5-fold in mice that either lacked or overexpressed ABCA1, respectively. However, ABCA1 had no effect on the FCR of Aβ, suggesting that ABCA1 does not regulate Aβ metabolism in the brain. Conclusions Our SILK strategy represents a straightforward, cost-effective, and efficient method to measure the clearance of proteins in the mouse brain. We expect that

  14. Isotopic labeling for the understanding of the alteration of limestone used in built cultural heritage

    Science.gov (United States)

    Saheb, Mandana; Chabas, Anne; Mertz, Jean-Didier; Rozenbaum, Olivier; Verney-Carron, Aurélie

    2015-04-01

    This project belongs to a specific work aiming at developing isotopic tools to better understand the alteration of materials used in the built cultural heritage. It is focused on the study of the alteration of limestone used in the facades of historic buildings subject to atmospheric polluted environment. Actually in the elevated parts of the buildings, water as rainfall (runoff or wet deposition) or in vapor form (condensation or dry deposition) is the main agent of alteration. Thus, the rock/water interactions need to be well understood to propose adapted solution to better preserve the buildings. To identify the water transfer within the porous limestone and locate the reaction preferential sites, two isotopic tracers (D and 18O) are used to monitor the alteration solution (D) and locate the zones containing the secondary phases (18O). The Saint-Maximin limestone used in many monuments in the suburbs of Paris (France) as a building and restoration stone has been specifically studied. Pristine materials, stones from monuments (monuments in the Paris area) and samples altered in laboratory constitute the analytical corpus to compare different stages of alteration. In a first step the stones are characterized at different scales to identify the alteration pattern (SEM-EDS, Raman microspectrometry, XRD, rugosimetry) and study the water transfers (X-ray tomography, mercury porosimetry, imbibition kinetics). The samples are then altered in the laboratory by realistic and controlled wet or dry deposition using isotopically labeled solutions to locate the reaction zones by SIMS. The multiscale characterization of the alteration pattern has allowed proposing alteration mechanisms linked to the properties of the stones and their location inside the building. Moreover, the location of the reactive zones inside the materials determined by the isotopic experiments helps examining the role of the evolution of porosity and formation of alteration products within the material

  15. Method for separation and enrichment of isotopes

    International Nuclear Information System (INIS)

    Kakihana, H.; Miyamatsu, T.

    1977-01-01

    Boron or uranium isotopes can be chemically separated and enriched with high speed and with high separating efficiency by using weakly basic anion exchange fibers having a diameter of not more than 100 μ, an aspect ratio of at least 5 and an exchange capacity of at least 2 meq/g-dry fiber, which are packed in a column at a specific volume of 2.0 to 20.0 ml/g-dry fiber

  16. Double-label autoradiographic deoxyglucose method for sequential measurement of regional cerebral glucose utilization

    Energy Technology Data Exchange (ETDEWEB)

    Redies, C; Diksic, M; Evans, A C; Gjedde, A; Yamamoto, Y L

    1987-08-01

    A new double-label autoradiographic glucose analog method for the sequential measurement of altered regional cerebral metabolic rates for glucose in the same animal is presented. This method is based on the sequential injection of two boluses of glucose tracer labeled with two different isotopes (short-lived /sup 18/F and long-lived /sup 3/H, respectively). An operational equation is derived which allows the determination of glucose utilization for the time period before the injection of the second tracer; this equation corrects for accumulation and loss of the first tracer from the metabolic pool occurring after the injection of the second tracer. An error analysis of this operational equation is performed. The double-label deoxyglucose method is validated in the primary somatosensory (''barrel'') cortex of the anesthetized rat. Two different rows of whiskers were stimulated sequentially in each rat; the two periods of stimulation were each preceded by an injection of glucose tracer. After decapitation, dried brain slices were first exposed, in direct contact, to standard X-ray film and then to uncoated, ''tritium-sensitive'' film. Results show that the double-label deoxyglucose method proposed in this paper allows the quantification and complete separation of glucose utilization patterns elicited by two different stimulations sequentially applied in the same animal.

  17. An isotope method for the measurement of creaming

    International Nuclear Information System (INIS)

    Wiechen, A.; Heine, K.

    1974-01-01

    The principle of a method is described which allows the course of creaming to be recorded continously and automatically by means of isotopes. Without affecting the colloidal system of milk, an isotope is added in hydrosoluble form and with only small amounts of carriers. A small detector sensitive to the respective radiation of the isotope is used to measure the decrease of counting rate on the head of the creaming cylinder, the decrease of rate being due to the effective recess of the radiation source and to the absorption of rays in the cream layer. The choice of the isotope, i.e. kind and energy of its radiation, and of the detector allows to adapt the sensitivity of the method to the rate of creaming. The method described appears to be superior to those techniques in which sedimentation balances are used; it could therefore supply useful information in research work on the process of creaming. (orig.) [de

  18. A convenient method to synthesize specifically labelled cholesterol with tritium

    International Nuclear Information System (INIS)

    Malik, S.; Kenny, M.; Ahmad, S.; Washington Univ., Seattle, WA

    1992-01-01

    A simple method is described to label cholesterol with tritium. Cholesterol was first oxidized to 5-cholesten-3-one which was then purified by HPLC. Its structure was established by electron impact (EI) mass spectrometry and 1 H-NMR spectroscopy. The ketone was reduced with NaB 3 H 4 to give specifically labelled cholesterol (C-3 3 H) at low specific activity. (author)

  19. Low energy methods of molecular laser isotope separation

    International Nuclear Information System (INIS)

    Makarov, G N

    2015-01-01

    Of the many proposals to date for laser-assisted isotope separation methods, isotope-selective infrared (IR) multiphoton dissociation (MPD) of molecules has been the most fully developed. This concept served as the basis for the development and operation of the carbon isotope separation facility in Kaliningrad, Russia. The extension of this method to heavy elements, including uranium, is hindered by, among other factors, the high power consumption and the lack of high-efficiency high-power laser systems. In this connection, research and development covering low energy methods for the laser separation of isotopes (including those of heavy atoms) is currently in high demand. This paper reviews approaches to the realization of IR-laser-induced isotope-selective processes, some of which are potentially the basis on which low-energy methods for molecular laser isotope separation can be developed. The basic physics and chemistry, application potential, and strengths and weaknesses of these approaches are discussed. Potentially promising alternatives to the title methods are examined. (reviews of topical problems)

  20. Determining synthesis rates of individual proteins in zebrafish (Danio rerio) with low levels of a stable isotope labelled amino acid.

    Science.gov (United States)

    Geary, Bethany; Magee, Kieran; Cash, Phillip; Young, Iain S; Whitfield, Phillip D; Doherty, Mary K

    2016-05-01

    The zebrafish is a powerful model organism for the analysis of human cardiovascular development and disease. Understanding these processes at the protein level not only requires changes in protein concentration to be determined but also the rate at which these changes occur on a protein-by-protein basis. The ability to measure protein synthesis and degradation rates on a proteome-wide scale, using stable isotope labelling in conjunction with mass spectrometry is now a well-established experimental approach. With the advent of more selective and sensitive mass spectrometers, it is possible to accurately measure lower levels of stable isotope incorporation, even when sample is limited. In order to challenge the sensitivity of this approach, we successfully determined the synthesis rates of over 600 proteins from the cardiac muscle of the zebrafish using a diet where either 30% or 50% of the L-leucine was replaced with a stable isotope labelled analogue ([(2) H7 ]L-leucine]. It was possible to extract sufficient protein from individual zebrafish hearts to determine the incorporation rate of the label into hundreds of proteins simultaneously, with the two labelling regimens showing a good correlation of synthesis rates. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Validation of 13CO2 breath analysis as a measurement of demethylation of stable isotope labeled aminopyrine in man

    International Nuclear Information System (INIS)

    Schneider, J.F.; Schoeller, D.A.; Nemchausky, B.; Bayer, J.L.; Klein, P.

    1978-01-01

    Interval sampling of expired breath as a simple, non-invasive assessment of the effect of liver disease upon hepatic microsomal drug metabolism, has been demonstrated with [ 14 C] dimethylaminoantipyrine (aminopyrine). In order to eliminate radiation risk the authors have validated the use of aminopyrine labeled with the stable, non-radioactive isotope 13 C. Simultaneous oral administration of both [ 14 C]- and [ 13 C] aminopyrine to five adult subjects without liver disease as well as five patients with known liver disease, resulted in the excretion of label at nearly identical rates in both individual time collections (r=0.94) as well as cumulative excretion for three hours (r=0.97). An oral dose of 2-mg/kg of [ 13 C) aminopyrine resulted in rates of production of 13 CO 2 significantly greater than baseline variations in 13 CO 2 production in the fasting, resting subject. Measurements of a single peak value at one half hour correlated closely with the determination of cumulative appearance over three hours (r=0.96). A consistent reproducible increase in the peak production of 13 CO 2 was observed when five patients received phenobarbital. Stable isotope labeled aminopyrine may be used to detect the effects of disease and treatment upon hepatic N-demethylation activity in human subjects without incurring any risk from radiation. Furthermore, the availability of another isotopic carbon label should make possible the study of direct drug-drug interaction utilizing CO 2 analysis. (Auth.)

  2. Isotope decay equations solved by means of a recursive method

    International Nuclear Information System (INIS)

    Grant, Carlos

    2009-01-01

    The isotope decay equations have been solved using forward finite differences taking small time steps, among other methods. This is the case of the cell code WIMS, where it is assumed that concentrations of all fissionable isotopes remain constant during the integration interval among other simplifications. Even when the problem could be solved running through a logical tree, all algorithms used for resolution of these equations used an iterative programming formulation. That happened because nearly all computer languages used up to a recent past by the scientific programmers did not support recursion, such as the case of the old versions of FORTRAN or BASIC. Nowadays also an integral form of the depletion equations is used in Monte Carlo simulation. In this paper we propose another programming solution using a recursive algorithm, running through all descendants of each isotope and adding their contributions to all isotopes in each generation. The only assumption made for this solution is that fluxes remain constant during the whole time step. Recursive process is interrupted when a stable isotope was attained or the calculated contributions are smaller than a given precision. These algorithms can be solved by means an exact analytic method that can have some problems when circular loops appear for isotopes with alpha decay, and a more general polynomial method. Both methods are shown. (author)

  3. Inter-labeler and intra-labeler variability of condition severity classification models using active and passive learning methods.

    Science.gov (United States)

    Nissim, Nir; Shahar, Yuval; Elovici, Yuval; Hripcsak, George; Moskovitch, Robert

    2017-09-01

    Labeling instances by domain experts for classification is often time consuming and expensive. To reduce such labeling efforts, we had proposed the application of active learning (AL) methods, introduced our CAESAR-ALE framework for classifying the severity of clinical conditions, and shown its significant reduction of labeling efforts. The use of any of three AL methods (one well known [SVM-Margin], and two that we introduced [Exploitation and Combination_XA]) significantly reduced (by 48% to 64%) condition labeling efforts, compared to standard passive (random instance-selection) SVM learning. Furthermore, our new AL methods achieved maximal accuracy using 12% fewer labeled cases than the SVM-Margin AL method. However, because labelers have varying levels of expertise, a major issue associated with learning methods, and AL methods in particular, is how to best to use the labeling provided by a committee of labelers. First, we wanted to know, based on the labelers' learning curves, whether using AL methods (versus standard passive learning methods) has an effect on the Intra-labeler variability (within the learning curve of each labeler) and inter-labeler variability (among the learning curves of different labelers). Then, we wanted to examine the effect of learning (either passively or actively) from the labels created by the majority consensus of a group of labelers. We used our CAESAR-ALE framework for classifying the severity of clinical conditions, the three AL methods and the passive learning method, as mentioned above, to induce the classifications models. We used a dataset of 516 clinical conditions and their severity labeling, represented by features aggregated from the medical records of 1.9 million patients treated at Columbia University Medical Center. We analyzed the variance of the classification performance within (intra-labeler), and especially among (inter-labeler) the classification models that were induced by using the labels provided by seven

  4. Voltammetry coupled to mass spectrometry in the presence of isotope {sup 18}O labeled water for the prediction of oxidative transformation pathways of activated aromatic ethers: Acebutolol

    Energy Technology Data Exchange (ETDEWEB)

    Bussy, Ugo; Tea, Illa [LUNAM Université de Nantes, CNRS, Chimie et Interdisciplinarité: Synthèse, Analyse et Modélisation (CEISAM), UMR 6230, 2 rue de la Houssinière, BP 92208, F-44322 Nantes cedex 3 (France); Ferchaud-Roucher, Véronique; Krempf, Michel [Université de Nantes, Plateforme Spectrométrie de Masse, CRNH, SFR Santé F. Bonamy, Institut du Thorax, UMR S1087, IRT-UN, BP 70721, 8 Quai Moncousu, 44007 Nantes cedex 1 (France); Silvestre, Virginie; Galland, Nicolas [LUNAM Université de Nantes, CNRS, Chimie et Interdisciplinarité: Synthèse, Analyse et Modélisation (CEISAM), UMR 6230, 2 rue de la Houssinière, BP 92208, F-44322 Nantes cedex 3 (France); Jacquemin, Denis [LUNAM Université de Nantes, CNRS, Chimie et Interdisciplinarité: Synthèse, Analyse et Modélisation (CEISAM), UMR 6230, 2 rue de la Houssinière, BP 92208, F-44322 Nantes cedex 3 (France); Institut Universitaire de France, 103, Boulevard Saint-Michel, 75005 Cedex 5 France (France); Andresen-Bergström, Moa; Jurva, Ulrik [CVGI iMed DMPK, AstraZeneca R and D Mölndal, Mölndal (Sweden); and others

    2013-01-31

    Highlights: ► Voltammetry coupled to mass spectrometry method as a useful tool for on-line predictions of electrochemical transformations. ► Evidence of the O-dealkoxylation reaction pathway of acebutolol in the presence of labeled water. ► New approach for on line EC-MS applications. -- Abstract: The coupling between an electrochemical cell (EC) and a mass spectrometer (MS) is a useful screening tool (EC-MS) to study the oxidative transformation pathways of various electroactive species. For that purpose, we showed that the EC-MS method, carried out in the presence and absence of isotope {sup 18}O labeled water leads not only to a fast identification of oxidation products but also leads to a fast elucidation of the mechanism pathway reaction. We examined herein the case of the electrochemical hydrolysis of activated aromatic ether. Acebutolol (β-blockers) was selected herein as model of activated aromatic ether, and its electrochemical oxidation was examined in both the presence and absence of isotope {sup 18}O labeled water. To elucidate electrochemical hydrolysis pathway reaction: O-dealkylation or O-dealkoxylation, our approach was used to prove its applicability. The electrochemical oxidation mechanism was then elucidated showing an O-dealkoxylation reaction. In addition, density functional theory (DFT) calculations fully support the experimental conclusions.

  5. Isotopic methods of investigations of hydrodynamics in installations for hydrometallurgic processing of copper concentrates

    International Nuclear Information System (INIS)

    Stsheletski, M.; Urban'ski, T.S.

    1979-01-01

    Isotope methods have been presented of investigations of hydrodynamics of liquid and solid phases in pilot-scale installations for hydrometallurgic processing of copper concentrates: in the column installation for leaching; in the tubular reactor for copper reduction by hydrogen and in the installation for crystallization of magnesium sulphate under pressure conditions. The column leaching installation has been tested by pulse injection of two radioisotope indicators simultaneously. The copper concentrate was labelled by sorbed colloidal gold-198 and the liquid phase was labelled by solution of sodium chloride containing sodium-24. Measurements of the radiation intensities were registered by scintillation counters, working with a single-channel amplitude analyzer, integrators and counting rate monitors. According to the distribution of time of stay, by the moments method, number of powers of ideal intermixings in the cell model of flow of the both phases. In the tubular reactor process of copper reduction by hydrogen is going on in three phases: gas-liquid-solig phase. Hydrodynamic investigations in this instalation was done in the presence of air, water and copper powder. Water was labelled by sodium-24 and copper powder by copper-64. Changes of intensity of radiation were measured by scintillation counters, located along the installation and were registered by multichannel amplitude analyzer. Peckle number and longitudal dispersion factor were determined. In investigations of the solid phase hydrodynamics during crystallization of magnesium sulphate under elevated temperature and high pressure, as an indicator, isotope gold-198 has been used, by which crystalls of the solid phase were labelled, and isotope sodium-24 was added to a liquid. Simultaneousely were measured and registered by a single-channel analyzer intensity of radiation of both indicators. Mean time of stay and parameters of a mathematical model of the phases flow in this installation were

  6. Quantitative proteomics by amino acid labeling in C. elegans

    DEFF Research Database (Denmark)

    Fredens, Julius; Engholm-Keller, Kasper; Giessing, Anders

    2011-01-01

    We demonstrate labeling of Caenorhabditis elegans with heavy isotope-labeled lysine by feeding them with heavy isotope-labeled Escherichia coli. Using heavy isotope-labeled worms and quantitative proteomics methods, we identified several proteins that are regulated in response to loss or RNAi-med......-mediated knockdown of the nuclear hormone receptor 49 in C. elegans. The combined use of quantitative proteomics and selective gene knockdown is a powerful tool for C. elegans biology.......We demonstrate labeling of Caenorhabditis elegans with heavy isotope-labeled lysine by feeding them with heavy isotope-labeled Escherichia coli. Using heavy isotope-labeled worms and quantitative proteomics methods, we identified several proteins that are regulated in response to loss or RNAi...

  7. Energy expenditures of plasma method of isotope separation

    International Nuclear Information System (INIS)

    Karchevskij, A.I.; Potanin, E.P.

    1986-01-01

    The estimations are performed of specific energy expenditares in isotope separation of binary mixtures in different plasma systems with weak medium ionization (plasma centrifuge, gas discharge system with travelling magnetic field, direct current discharge). Potential advantages of plasma centrifuge over other gas discharge facilities are pointed out. The comparison of specific energy expenditure values in case of using plasma and conventional methods of isotope separation is carried out

  8. Method and apparatus for controlled condensation isotope separation

    International Nuclear Information System (INIS)

    Sullivan, J.A.; Lee, J.T. Jr.; Kim, K.C.

    1981-01-01

    The invention provides a method for producing controlled homogeneous condensation of a molecular feed gas containing several isotopes. The feed gas flows at supersonic rates through an expansion nozzle under conditions at which the gas would normally condense. The gas is irradiated with laser radiation of a wavelength that selectively excites those molecules in the feed gas that contain a particular isotope, thus preventing their condensation. Condensate particles may be aerodynamically separated from the flowing gas stream

  9. Isotopically modified compounds

    International Nuclear Information System (INIS)

    Kuruc, J.

    2009-01-01

    In this chapter the nomenclature of isotopically modified compounds in Slovak language is described. This chapter consists of following parts: (1) Isotopically substituted compounds; (2) Specifically isotopically labelled compounds; (3) Selectively isotopically labelled compounds; (4) Non-selectively isotopically labelled compounds; (5) Isotopically deficient compounds.

  10. Labelling of long chain fatty acids by non isotopic nucleophilic halogen exchange

    International Nuclear Information System (INIS)

    Hallaba, E.; Al-Suhybani, A.A.; Zaki, F.S.

    1985-01-01

    The parameters of two exchange methods for preparing pure 97% labelled 17-Br-HDA in acetone and in benzene with dry NaI in a closed system are described. In aprotic solvents the need for a phase transfer catalyst up to 50 μg is necessary to dissolve the dry NaI. The use of aqueous medium in the exchange is totally prohibited. Energies of activation are calculated for both reactions. (author)

  11. Application of SAIL phenylalanine and tyrosine with alternative isotope-labeling patterns for protein structure determination

    Energy Technology Data Exchange (ETDEWEB)

    Takeda, Mitsuhiro [Nagoya University, Structural Biology Research Center, Graduate School of Science (Japan); Ono, Akira M.; Terauchi, Tsutomu [SAIL Technologies Co., Inc. (Japan); Kainosho, Masatsune, E-mail: kainosho@nmr.chem.metro-u.ac.j [Nagoya University, Structural Biology Research Center, Graduate School of Science (Japan)

    2010-01-15

    The extensive collection of NOE constraint data involving the aromatic ring signals is essential for accurate protein structure determination, although it is often hampered in practice by the pervasive signal overlapping and tight spin couplings for aromatic rings. We have prepared various types of stereo-array isotope labeled phenylalanines ({epsilon}- and {zeta}-SAIL Phe) and tyrosine ({epsilon}-SAIL Tyr) to overcome these problems (Torizawa et al. 2005), and proven that these SAIL amino acids provide dramatic spectral simplification and sensitivity enhancement for the aromatic ring NMR signals. In addition to these SAIL aromatic amino acids, we recently synthesized {delta}-SAIL Phe and {delta}-SAIL Tyr, which allow us to observe and assign {delta}-{sup 13}C/{sup 1}H signals very efficiently. Each of the various types of SAIL Phe and SAIL Tyr yields well-resolved resonances for the {delta}-, {epsilon}- or {zeta}-{sup 13}C/{sup 1}H signals, respectively, which can readily be assigned by simple and robust pulse sequences. Since the {delta}-, {epsilon}-, and {zeta}-proton signals of Phe/Tyr residues give rise to complementary NOE constraints, the concomitant use of various types of SAIL-Phe and SAIL-Tyr would generate more accurate protein structures, as compared to those obtained by using conventional uniformly {sup 13}C, {sup 15}N-double labeled proteins. We illustrated this with the case of an 18.2 kDa protein, Escherichia coli peptidyl-prolyl cis-trans isomerase b (EPPIb), and concluded that the combined use of {zeta}-SAIL Phe and {epsilon}-SAIL Tyr would be practically the best choice for protein structural determinations.

  12. Application of SAIL phenylalanine and tyrosine with alternative isotope-labeling patterns for protein structure determination.

    Science.gov (United States)

    Takeda, Mitsuhiro; Ono, Akira M; Terauchi, Tsutomu; Kainosho, Masatsune

    2010-01-01

    The extensive collection of NOE constraint data involving the aromatic ring signals is essential for accurate protein structure determination, although it is often hampered in practice by the pervasive signal overlapping and tight spin couplings for aromatic rings. We have prepared various types of stereo-array isotope labeled phenylalanines (epsilon- and zeta-SAIL Phe) and tyrosine (epsilon-SAIL Tyr) to overcome these problems (Torizawa et al. 2005), and proven that these SAIL amino acids provide dramatic spectral simplification and sensitivity enhancement for the aromatic ring NMR signals. In addition to these SAIL aromatic amino acids, we recently synthesized delta-SAIL Phe and delta-SAIL Tyr, which allow us to observe and assign delta-(13)C/(1)H signals very efficiently. Each of the various types of SAIL Phe and SAIL Tyr yields well-resolved resonances for the delta-, epsilon- or zeta-(13)C/(1)H signals, respectively, which can readily be assigned by simple and robust pulse sequences. Since the delta-, epsilon-, and zeta-proton signals of Phe/Tyr residues give rise to complementary NOE constraints, the concomitant use of various types of SAIL-Phe and SAIL-Tyr would generate more accurate protein structures, as compared to those obtained by using conventional uniformly (13)C, (15)N-double labeled proteins. We illustrated this with the case of an 18.2 kDa protein, Escherichia coli peptidyl-prolyl cis-trans isomerase b (EPPIb), and concluded that the combined use of zeta-SAIL Phe and epsilon-SAIL Tyr would be practically the best choice for protein structural determinations.

  13. Application of SAIL phenylalanine and tyrosine with alternative isotope-labeling patterns for protein structure determination

    International Nuclear Information System (INIS)

    Takeda, Mitsuhiro; Ono, Akira M.; Terauchi, Tsutomu; Kainosho, Masatsune

    2010-01-01

    The extensive collection of NOE constraint data involving the aromatic ring signals is essential for accurate protein structure determination, although it is often hampered in practice by the pervasive signal overlapping and tight spin couplings for aromatic rings. We have prepared various types of stereo-array isotope labeled phenylalanines (ε- and ζ-SAIL Phe) and tyrosine (ε-SAIL Tyr) to overcome these problems (Torizawa et al. 2005), and proven that these SAIL amino acids provide dramatic spectral simplification and sensitivity enhancement for the aromatic ring NMR signals. In addition to these SAIL aromatic amino acids, we recently synthesized δ-SAIL Phe and δ-SAIL Tyr, which allow us to observe and assign δ- 13 C/ 1 H signals very efficiently. Each of the various types of SAIL Phe and SAIL Tyr yields well-resolved resonances for the δ-, ε- or ζ- 13 C/ 1 H signals, respectively, which can readily be assigned by simple and robust pulse sequences. Since the δ-, ε-, and ζ-proton signals of Phe/Tyr residues give rise to complementary NOE constraints, the concomitant use of various types of SAIL-Phe and SAIL-Tyr would generate more accurate protein structures, as compared to those obtained by using conventional uniformly 13 C, 15 N-double labeled proteins. We illustrated this with the case of an 18.2 kDa protein, Escherichia coli peptidyl-prolyl cis-trans isomerase b (EPPIb), and concluded that the combined use of ζ-SAIL Phe and ε-SAIL Tyr would be practically the best choice for protein structural determinations.

  14. Inter-Labeler and Intra-Labeler Variability of Condition Severity Classification Models Using Active and Passive Learning Methods

    Science.gov (United States)

    Nissim, Nir; Shahar, Yuval; Boland, Mary Regina; Tatonetti, Nicholas P; Elovici, Yuval; Hripcsak, George; Moskovitch, Robert

    2018-01-01

    Background and Objectives Labeling instances by domain experts for classification is often time consuming and expensive. To reduce such labeling efforts, we had proposed the application of active learning (AL) methods, introduced our CAESAR-ALE framework for classifying the severity of clinical conditions, and shown its significant reduction of labeling efforts. The use of any of three AL methods (one well known [SVM-Margin], and two that we introduced [Exploitation and Combination_XA]) significantly reduced (by 48% to 64%) condition labeling efforts, compared to standard passive (random instance-selection) SVM learning. Furthermore, our new AL methods achieved maximal accuracy using 12% fewer labeled cases than the SVM-Margin AL method. However, because labelers have varying levels of expertise, a major issue associated with learning methods, and AL methods in particular, is how to best to use the labeling provided by a committee of labelers. First, we wanted to know, based on the labelers’ learning curves, whether using AL methods (versus standard passive learning methods) has an effect on the Intra-labeler variability (within the learning curve of each labeler) and inter-labeler variability (among the learning curves of different labelers). Then, we wanted to examine the effect of learning (either passively or actively) from the labels created by the majority consensus of a group of labelers. Methods We used our CAESAR-ALE framework for classifying the severity of clinical conditions, the three AL methods and the passive learning method, as mentioned above, to induce the classifications models. We used a dataset of 516 clinical conditions and their severity labeling, represented by features aggregated from the medical records of 1.9 million patients treated at Columbia University Medical Center. We analyzed the variance of the classification performance within (intra-labeler), and especially among (inter-labeler) the classification models that were induced by

  15. Stable isotope applications in biomolecular structure and mechanisms. A meeting to bring together producers and users of stable-isotope-labeled compounds to assess current and future needs

    International Nuclear Information System (INIS)

    Trewhella, J.; Cross, T.A.; Unkefer, C.J.

    1994-12-01

    Knowledge of biomolecular structure is a prerequisite for understanding biomolecular function, and stable isotopes play an increasingly important role in structure determination of biological molecules. The first Conference on Stable Isotope Applications in Biomolecular Structure and Mechanisms was held in Santa Fe, New Mexico, March 27--31, 1994. More than 120 participants from 8 countries and 44 institutions reviewed significant developments, discussed the most promising applications for stable isotopes, and addressed future needs and challenges. Participants focused on applications of stable isotopes for studies of the structure and function of proteins, peptides, RNA, and DNA. Recent advances in NMR techniques neutron scattering, EPR, and vibrational spectroscopy were highlighted in addition to the production and synthesis of labeled compounds. This volume includes invited speaker and poster presentations as well as a set of reports from discussion panels that focused on the needs of the scientific community and the potential roles of private industry, the National Stable Isotope Resource, and the National High Magnetic Field Laboratory in serving those needs. This is the leading abstract. Individual papers are processed separately for the database

  16. Stable isotope applications in biomolecular structure and mechanisms. A meeting to bring together producers and users of stable-isotope-labeled compounds to assess current and future needs

    Energy Technology Data Exchange (ETDEWEB)

    Trewhella, J.; Cross, T.A.; Unkefer, C.J. [eds.

    1994-12-01

    Knowledge of biomolecular structure is a prerequisite for understanding biomolecular function, and stable isotopes play an increasingly important role in structure determination of biological molecules. The first Conference on Stable Isotope Applications in Biomolecular Structure and Mechanisms was held in Santa Fe, New Mexico, March 27--31, 1994. More than 120 participants from 8 countries and 44 institutions reviewed significant developments, discussed the most promising applications for stable isotopes, and addressed future needs and challenges. Participants focused on applications of stable isotopes for studies of the structure and function of proteins, peptides, RNA, and DNA. Recent advances in NMR techniques neutron scattering, EPR, and vibrational spectroscopy were highlighted in addition to the production and synthesis of labeled compounds. This volume includes invited speaker and poster presentations as well as a set of reports from discussion panels that focused on the needs of the scientific community and the potential roles of private industry, the National Stable Isotope Resource, and the National High Magnetic Field Laboratory in serving those needs. This is the leading abstract. Individual papers are processed separately for the database.

  17. Isotope label-aided mass spectrometry reveals the influence of environmental factors on metabolism in single eggs of fruit fly.

    Directory of Open Access Journals (Sweden)

    Te-Wei Tseng

    Full Text Available In order to investigate the influence of light/dark cycle on the biosynthesis of metabolites during oogenesis, here we demonstrate a simple experimental protocol which combines in-vivo isotopic labeling of primary metabolites with mass spectrometric analysis of single eggs of fruit fly (Drosophila melanogaster. First, fruit flies were adapted to light/dark cycle using artificial white light. Second, female flies were incubated with an isotopically labeled sugar ((13C(6-glucose for 12 h--either during the circadian day or the circadian night, at light or at dark. Third, eggs were obtained from the incubated female flies, and analyzed individually by matrix-assisted laser desorption/ionization (MALDI mass spectrometry (MS: this yielded information about the extent of labeling with carbon-13. Since the incorporation of carbon-13 to uridine diphosphate glucose (UDP-glucose in fruit fly eggs is very fast, the labeling of this metabolite was used as an indicator of the biosynthesis of metabolites flies/eggs during 12-h periods, which correspond to circadian day or circadian night. The results reveal that once the flies adapted to the 12-h-light/12-h-dark cycle, the incorporation of carbon-13 to UDP-glucose present in fruit fly eggs was not markedly altered by an acute perturbation to this cycle. This effect may be due to a relationship between biosynthesis of primary metabolites in developing eggs and an alteration to the intake of the labeled substrate - possibly related to the change of the feeding habit. Overall, the study shows the possibility of using MALDI-MS in conjunction with isotopic labeling of small metazoans to unravel the influence of environmental cues on primary metabolism.

  18. Auto-inducing media for uniform isotope labeling of proteins with 15N, 13C and 2H

    International Nuclear Information System (INIS)

    Guthertz, Nicolas; Klopp, Julia; Winterhalter, Aurélie; Fernández, César; Gossert, Alvar D.

    2015-01-01

    Auto-inducing media for protein expression offer many advantages like robust reproducibility, high yields of soluble protein and much reduced workload. Here, an auto-inducing medium for uniform isotope labelling of proteins with 15 N, 13 C and/or 2 H in E. coli is presented. So far, auto-inducing media have not found widespread application in the NMR field, because of the prohibitively high cost of labeled lactose, which is an essential ingredient of such media. Here, we propose using lactose that is only selectively labeled on the glucose moiety. It can be synthesized from inexpensive and readily available substrates: labeled glucose and unlabeled activated galactose. With this approach, uniformly isotope labeled proteins were expressed in unattended auto-inducing cultures with incorporation of 13 C, 15 N of 96.6 % and 2 H, 15 N of 98.8 %. With the present protocol, the NMR community could profit from the many advantages that auto-inducing media offer

  19. Stable isotope labeling, in vivo, of D- and L-tryptophan pools in lemna gibba and the low incorporation of label into indole-3-acetic acid

    International Nuclear Information System (INIS)

    Baldi, B.G.; Maher, B.R.; Slovin, J.P.; Cohen, J.D.

    1991-01-01

    The authors present evidence that the role of tryptophan and other potential intermediates in the pathways that could lead to indole derivatives needs to be reexamined. Two lines of Lemna gibba were tested for uptake of [ 15 N-indole]-labeled tryptophan isomers and incorporation of that label into free indole-3-acetic acid (IAA). Both lines required levels of L-[ 15 N]tryptophan 2 to 3 orders of magnitude over endogenous levels in order to obtain measurable incorporation of label into IAA. Labeled L-tryptophan was extractable from plant tissue after feeding and showed no measurable isomerization into D-tryptophan. D-[ 15 N]trytophan supplied to Lemna at rates of approximately 400 times excess of endogenous D-tryptophan levels (to yield an isotopic enrichment equal to that which allowed detection of the incorporation of L-tryptophan into IAA), did not result in measurable incorporation of label into free IAA. These results demonstrate that L-tryptophan is a more direct precursor to IAA than the D isomer and suggest (a) that the availability of tryptophan in vivo is not a limiting factor in the biosynthesis of IAA, thus implying that other regulatory mechanisms are in operation and (b) that L-tryptophan also may not be a primary precursor to IAA in plants

  20. Application of isotope-labelled compounds in the study of the chemical stability of pesticides

    International Nuclear Information System (INIS)

    Roesseler, M.; Luther, D.; Abendroth, H.C.; Koch, H.

    1980-01-01

    The user of pesticides requires specific biological modes of action from the corresponding commercial products. Impurities and degradation products may cause uncontrollable toxicological reactions. Profound knowledge of the chemical stability of the effective substance in question and its formulations under storage conditions as well as under those of analytical sample preparation and detection is required. Radioisotope labelled effective substances dimethoate and 1-butyl-amino-cyclohexane-phosphonic acid dibutyl ester are used to study storage stability of the pure effective substance and its formulations; effects of selected impurities, such as technical by-products, moisture or water content, binding or carrier materials, organic solvents, chemical stabilizers and other formulation components on storage properties; temperature dependence of storage stability; selection of suitable analytical techniques for quantitative determination of the effective substance without interference effects from any by-product; reduction of the necessary analytical expense; disclosure of sources of error in the application of usual analytical techniques; improvement of possibilities of an immediate and clearer discrimination between types and amounts of compounds in a chemical system consisting of one pesticide and its degradation or reaction products at the beginning and at the end of an experimental or reaction period. Radiochemical analytical techniques, such as radio thin-layer chromatography (also combined with liquid scintillation counting), radio gas chromatography, autoradiography and isotope dilution analysis were used. Results are discussed, especially of experiments on dimethoate and its technical by-products

  1. Uniform isotope labeling of a eukaryotic seven-transmembrane helical protein in yeast enables high-resolution solid-state NMR studies in the lipid environment

    International Nuclear Information System (INIS)

    Fan Ying; Shi Lichi; Ladizhansky, Vladimir; Brown, Leonid S.

    2011-01-01

    Overexpression of isotope-labeled multi-spanning eukaryotic membrane proteins for structural NMR studies is often challenging. On the one hand, difficulties with achieving proper folding, membrane insertion, and native-like post-translational modifications frequently disqualify bacterial expression systems. On the other hand, eukaryotic cell cultures can be prohibitively expensive. One of the viable alternatives, successfully used for producing proteins for solution NMR studies, is yeast expression systems, particularly Pichia pastoris. We report on successful implementation and optimization of isotope labeling protocols, previously used for soluble secreted proteins, to produce homogeneous samples of a eukaryotic seven-transmembrane helical protein, rhodopsin from Leptosphaeria maculans. Even in shake-flask cultures, yields exceeded 5 mg of purified uniformly 13 C, 15 N-labeled protein per liter of culture. The protein was stable (at least several weeks at 5°C) and functionally active upon reconstitution into lipid membranes at high protein-to-lipid ratio required for solid-state NMR. The samples gave high-resolution 13 C and 15 N solid-state magic angle spinning NMR spectra, amenable to a detailed structural analysis. We believe that similar protocols can be adopted for challenging mammalian targets, which often resist characterization by other structural methods.

  2. iMS2Flux – a high–throughput processing tool for stable isotope labeled mass spectrometric data used for metabolic flux analysis

    Directory of Open Access Journals (Sweden)

    Poskar C Hart

    2012-11-01

    Full Text Available Abstract Background Metabolic flux analysis has become an established method in systems biology and functional genomics. The most common approach for determining intracellular metabolic fluxes is to utilize mass spectrometry in combination with stable isotope labeling experiments. However, before the mass spectrometric data can be used it has to be corrected for biases caused by naturally occurring stable isotopes, by the analytical technique(s employed, or by the biological sample itself. Finally the MS data and the labeling information it contains have to be assembled into a data format usable by flux analysis software (of which several dedicated packages exist. Currently the processing of mass spectrometric data is time-consuming and error-prone requiring peak by peak cut-and-paste analysis and manual curation. In order to facilitate high-throughput metabolic flux analysis, the automation of multiple steps in the analytical workflow is necessary. Results Here we describe iMS2Flux, software developed to automate, standardize and connect the data flow between mass spectrometric measurements and flux analysis programs. This tool streamlines the transfer of data from extraction via correction tools to 13C-Flux software by processing MS data from stable isotope labeling experiments. It allows the correction of large and heterogeneous MS datasets for the presence of naturally occurring stable isotopes, initial biomass and several mass spectrometry effects. Before and after data correction, several checks can be performed to ensure accurate data. The corrected data may be returned in a variety of formats including those used by metabolic flux analysis software such as 13CFLUX, OpenFLUX and 13CFLUX2. Conclusion iMS2Flux is a versatile, easy to use tool for the automated processing of mass spectrometric data containing isotope labeling information. It represents the core framework for a standardized workflow and data processing. Due to its flexibility

  3. Applications of isotopic labeling to the study of Friedel-Crafts reactions

    International Nuclear Information System (INIS)

    Roberts, R.M.; Gibson, T.L.

    1980-01-01

    A number of Friedel-Crafts processes that have been examined by the use of isotopic methods are considered: (1) the interaction of a Lewis acid with an acyl or alkyl halide; (2) the rearrangements of an alkylating species resulting from such an interaction, as observed in the arene products or in recovered starting materials; and (3) isomerizations of alkylation products which can occur in the presence of Lewis acids including (a) the alkylbenzene rearrangement (internal, of side chains), (b) disproportionation (intermolecular alkyl transfers), and (c) reorientations, which may be the results of either intra- or intermolecular processes. (Auth.)

  4. Compensation of matrix effects in gas chromatography-mass spectrometry analysis of pesticides using a combination of matrix matching and multiple isotopically labeled internal standards.

    Science.gov (United States)

    Tsuchiyama, Tomoyuki; Katsuhara, Miki; Nakajima, Masahiro

    2017-11-17

    In the multi-residue analysis of pesticides using GC-MS, the quantitative results are adversely affected by a phenomenon known as the matrix effect. Although the use of matrix-matched standards is considered to be one of the most practical solutions to this problem, complete removal of the matrix effect is difficult in complex food matrices owing to their inconsistency. As a result, residual matrix effects can introduce analytical errors. To compensate for residual matrix effects, we have developed a novel method that employs multiple isotopically labeled internal standards (ILIS). The matrix effects of ILIS and pesticides were evaluated in spiked matrix extracts of various agricultural commodities, and the obtained data were subjected to simple statistical analysis. Based on the similarities between the patterns of variation in the analytical response, a total of 32 isotopically labeled compounds were assigned to 338 pesticides as internal standards. It was found that by utilizing multiple ILIS, residual matrix effects could be effectively compensated. The developed method exhibited superior quantitative performance compared with the common single-internal-standard method. The proposed method is more feasible for regulatory purposes than that using only predetermined correction factors and is considered to be promising for practical applications. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Document clustering methods, document cluster label disambiguation methods, document clustering apparatuses, and articles of manufacture

    Science.gov (United States)

    Sanfilippo, Antonio [Richland, WA; Calapristi, Augustin J [West Richland, WA; Crow, Vernon L [Richland, WA; Hetzler, Elizabeth G [Kennewick, WA; Turner, Alan E [Kennewick, WA

    2009-12-22

    Document clustering methods, document cluster label disambiguation methods, document clustering apparatuses, and articles of manufacture are described. In one aspect, a document clustering method includes providing a document set comprising a plurality of documents, providing a cluster comprising a subset of the documents of the document set, using a plurality of terms of the documents, providing a cluster label indicative of subject matter content of the documents of the cluster, wherein the cluster label comprises a plurality of word senses, and selecting one of the word senses of the cluster label.

  6. Dual isotope study of iodine-125 and indium-111-labeled antibody in athymic mice

    Energy Technology Data Exchange (ETDEWEB)

    Carney, P.L.; Rogers, P.E.; Johnson, D.K. (Abbott Laboratories, Abbott Park, IL (USA))

    1989-03-01

    Monoclonal antibody B72.3 was coupled to a benzylisothiocyanate derivative of diethylenetriaminepentaacetic acid (DTPA). The maximum substitution achievable without loss of immunoreactivity was three DTPA groups per immunoglobulin molecule. The resulting conjugate was labeled with {sup 111}In by brief incubation with {sup 111}InCl{sub 3}, giving a mean radiochemical yield of {sup 111}In-labeled antibody of 96%. The ({sup 111}In)B72.3 preparation was mixed with an ({sup 125}I) B72.3 preparation, obtained by the chloramine-T method, and the mixture administered to athymic mice bearing subcutaneous LS174T colon carcinoma xenografts. There were no significant differences (p greater than 0.1) in the biodistributions of the two labels at 1, 2, 5, and 7 days postinjection. These results are contrasted with prior studies showing elevated levels of {sup 111}In in liver, spleen, and kidneys using B72.3-DTPA conjugates prepared via the bicyclic anhydride. It is concluded that protein cross-linking and/or the formation of unstable chelate sites in anhydride coupled conjugates underlie these disparities.

  7. Dual isotope study of iodine-125 and indium-111-labeled antibody in athymic mice

    International Nuclear Information System (INIS)

    Carney, P.L.; Rogers, P.E.; Johnson, D.K.

    1989-01-01

    Monoclonal antibody B72.3 was coupled to a benzylisothiocyanate derivative of diethylenetriaminepentaacetic acid (DTPA). The maximum substitution achievable without loss of immunoreactivity was three DTPA groups per immunoglobulin molecule. The resulting conjugate was labeled with 111 In by brief incubation with 111 InCl 3 , giving a mean radiochemical yield of 111 In-labeled antibody of 96%. The [ 111 In]B72.3 preparation was mixed with an [ 125 I] B72.3 preparation, obtained by the chloramine-T method, and the mixture administered to athymic mice bearing subcutaneous LS174T colon carcinoma xenografts. There were no significant differences (p greater than 0.1) in the biodistributions of the two labels at 1, 2, 5, and 7 days postinjection. These results are contrasted with prior studies showing elevated levels of 111 In in liver, spleen, and kidneys using B72.3-DTPA conjugates prepared via the bicyclic anhydride. It is concluded that protein cross-linking and/or the formation of unstable chelate sites in anhydride coupled conjugates underlie these disparities

  8. Hg stable isotope analysis by the double-spike method.

    Science.gov (United States)

    Mead, Chris; Johnson, Thomas M

    2010-06-01

    Recent publications suggest great potential for analysis of Hg stable isotope abundances to elucidate sources and/or chemical processes that control the environmental impact of mercury. We have developed a new MC-ICP-MS method for analysis of mercury isotope ratios using the double-spike approach, in which a solution containing enriched (196)Hg and (204)Hg is mixed with samples and provides a means to correct for instrumental mass bias and most isotopic fractionation that may occur during sample preparation and introduction into the instrument. Large amounts of isotopic fractionation induced by sample preparation and introduction into the instrument (e.g., by batch reactors) are corrected for. This may greatly enhance various Hg pre-concentration methods by correcting for minor fractionation that may occur during preparation and removing the need to demonstrate 100% recovery. Current precision, when ratios are normalized to the daily average, is 0.06 per thousand, 0.06 per thousand, 0.05 per thousand, and 0.05 per thousand (2sigma) for (202)Hg/(198)Hg, (201)Hg/(198)Hg, (200)Hg/(198)Hg, and (199)Hg/(198)Hg, respectively. This is slightly better than previously published methods. Additionally, this precision was attained despite the presence of large amounts of other Hg isotopes (e.g., 5.0% atom percent (198)Hg) in the spike solution; substantially better precision could be achieved if purer (196)Hg were used.

  9. Immunoreactivity of 125I-papain labelled by different methods

    International Nuclear Information System (INIS)

    Rauch, P.; Fukal, L.; Kas, J.; Tykva, R.

    1984-01-01

    Three different methods of papain iodination (with chloramine-T, lactoperoxidase and conjugation with Bolton-Hunter reagent) have been compared. The highest yield of 125 I-papain could be obtained using lactoperoxidase which enabled to achieve the highest immunoreactivity. 125 I-papain, labelled this way, is suitable for the radioimmunoassay of papain. (author)

  10. Sulfur Isotope Exchange between S-35 Labeled Inorganic Sulfur-Compounds in Anoxic Marine-Sediments

    DEFF Research Database (Denmark)

    FOSSING, H.; THODEANDERSEN, S.; JØRGENSEN, BB

    1992-01-01

    of isotope exchange, specific radioactivities of the reduced sulfur pools were poorly defined and could not be used to calculate their rates of formation. Such isotope exchange reactions between the reduced inorganic sulfur compounds will affect the stable isotope distribution and are expected to decrease...

  11. The application of high-resolution IR spectroscopy and isotope labeling for detailed investigation of TiO2/gas interface reactions

    Czech Academy of Sciences Publication Activity Database

    Civiš, Svatopluk; Ferus, Martin; Zukalová, Markéta; Kavan, Ladislav; Zelinger, Zdeněk

    2013-01-01

    Roč. 36, č. 1 (2013), s. 159-162 ISSN 0925-3467 R&D Projects: GA ČR GAP208/10/2302; GA ČR GA13-07724S Institutional support: RVO:61388955 Keywords : isotope exchange * titania * isotope labeling Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.075, year: 2013

  12. Stable isotope labeling by amino acids in cell culture, SILAC, as a simple and accurate approach to expression proteomics

    DEFF Research Database (Denmark)

    Ong, S.E.; Blagoev, B.; Kratchmarova, I.

    2002-01-01

    Quantitative proteomics has traditionally been performed by two-dimensional gel electrophoresis, but recently, mass spectrometric methods based on stable isotope quantitation have shown great promise for the simultaneous and automated identification and quantitation of complex protein mixtures. H...

  13. An isotope-dilution standard GC/MS/MS method for steroid hormones in water

    Science.gov (United States)

    Foreman, William T.; Gray, James L.; ReVello, Rhiannon C.; Lindley, Chris E.; Losche, Scott A.

    2013-01-01

    An isotope-dilution quantification method was developed for 20 natural and synthetic steroid hormones and additional compounds in filtered and unfiltered water. Deuterium- or carbon-13-labeled isotope-dilution standards (IDSs) are added to the water sample, which is passed through an octadecylsilyl solid-phase extraction (SPE) disk. Following extract cleanup using Florisil SPE, method compounds are converted to trimethylsilyl derivatives and analyzed by gas chromatography with tandem mass spectrometry. Validation matrices included reagent water, wastewater-affected surface water, and primary (no biological treatment) and secondary wastewater effluent. Overall method recovery for all analytes in these matrices averaged 100%; with overall relative standard deviation of 28%. Mean recoveries of the 20 individual analytes for spiked reagent-water samples prepared along with field samples analyzed in 2009–2010 ranged from 84–104%, with relative standard deviations of 6–36%. Detection levels estimated using ASTM International’s D6091–07 procedure range from 0.4 to 4 ng/L for 17 analytes. Higher censoring levels of 100 ng/L for bisphenol A and 200 ng/L for cholesterol and 3-beta-coprostanol are used to prevent bias and false positives associated with the presence of these analytes in blanks. Absolute method recoveries of the IDSs provide sample-specific performance information and guide data reporting. Careful selection of labeled compounds for use as IDSs is important because both inexact IDS-analyte matches and deuterium label loss affect an IDS’s ability to emulate analyte performance. Six IDS compounds initially tested and applied in this method exhibited deuterium loss and are not used in the final method.

  14. Site-specific Orientation of an α-helical Peptide Ovispirin-1 from Isotope Labeled SFG Spectroscopy

    Science.gov (United States)

    Ding, Bei; Laaser, Jennifer E.; Liu, Yuwei; Wang, Pengrui; Zanni, Martin T.; Chen, Zhan

    2013-01-01

    Sum-frequency generation (SFG) vibrational spectroscopy is often used to probe the backbone structures and orientations of polypeptides at surfaces. Using the ovispirin-1 polypeptide at the solid/liquid interface of polystyrene, we demonstrate for the first time that SFG can probe the polarization response of a single isotope labeled residue. To interpret the spectral intensities, we simulated the spectra using an excitonic Hamiltonian approach. We show that the polarization dependence of either the label or the unlabeled amide I band alone does not provide sufficient structural constraints to obtain both the tilt and the twist of the ovispirin helix at a solid/liquid interface, but that both can be determined from the polarization dependence of the complete spectrum. For ovispirin, the detailed analysis of the polarized SFG experimental data shows that the helix axis is tilted at roughly 138 degrees from the surface normal, and the transition dipole of the isotope labeled C=O group is tilted at 23 degrees from the surface normal, with the hydrophobic region facing the polystyrene surface. We further demonstrated that the Hamiltonian approach is able to address the coupling effect and the structural disorder. For comparison, we also collected the FTIR spectrum of ovispirin under similar conditions, which reveals the enhanced sensitivity of SFG for structural studies of single monolayer peptide surfaces. Our study provides insight into how structural and environmental effects appear in SFG spectra of the amide I band and establishes that SFG of isotope labeled peptides will be a powerful technique for elucidating secondary structures with residue-by-residue resolution. PMID:24228619

  15. Site-specific orientation of an α-helical peptide ovispirin-1 from isotope-labeled SFG spectroscopy.

    Science.gov (United States)

    Ding, Bei; Laaser, Jennifer E; Liu, Yuwei; Wang, Pengrui; Zanni, Martin T; Chen, Zhan

    2013-11-27

    Sum-frequency generation (SFG) vibrational spectroscopy is often used to probe the backbone structures and orientations of polypeptides at surfaces. Using the ovispirin-1 polypeptide at the solid/liquid interface of polystyrene, we demonstrate for the first time that SFG can probe the polarization response of a single-isotope-labeled residue. To interpret the spectral intensities, we simulated the spectra using an excitonic Hamiltonian approach. We show that the polarization dependence of either the label or the unlabeled amide I band alone does not provide sufficient structural constraints to obtain both the tilt and the twist of the ovispirin helix at a solid/liquid interface, but that both can be determined from the polarization dependence of the complete spectrum. For ovispirin, the detailed analysis of the polarized SFG experimental data shows that the helix axis is tilted at roughly 138° from the surface normal, and the transition dipole of the isotope-labeled C═O group is tilted at 23° from the surface normal, with the hydrophobic region facing the polystyrene surface. We further demonstrate that the Hamiltonian approach is able to address the coupling effect and the structural disorder. For comparison, we also collected the FTIR spectrum of ovispirin under similar conditions, which reveals the enhanced sensitivity of SFG for structural studies of single monolayer peptide surfaces. Our study provides insight into how structural and environmental effects appear in SFG spectra of the amide I band and establishes that SFG of isotope-labeled peptides will be a powerful technique for elucidating secondary structures with residue-by-residue resolution.

  16. Generator Coordinate Method Analysis of Xe and Ba Isotopes

    Science.gov (United States)

    Higashiyama, Koji; Yoshinaga, Naotaka; Teruya, Eri

    Nuclear structure of Xe and Ba isotopes is studied in terms of the quantum-number projected generator coordinate method (GCM). The GCM reproduces well the energy levels of high-spin states as well as low-lying states. The structure of the low-lying states is analyzed through the GCM wave functions.

  17. Overview on recent developments: alternative isotope production methods in Canada

    International Nuclear Information System (INIS)

    Huynh, K.

    2012-01-01

    The purpose of this paper is to provide an update on the Government of Canada's programs in alternative isotope production methods for securing supply of technetium 99m for Canadians. The supply disruptions of isotopes in 2007 and 2009/2010 caused by unplanned outages at AECL's National Research Universal (NRU) reactor highlighted the fragility of the supply chain that delivers medical isotopes, specifically Technetium 99m (Tc99m) to patients in Canada and globally. Tc99m, which is derived from its parent, molybdenum99 (Mo99) is the most widely used medical isotope for imaging, and accounts for 80 percent of nuclear medicine diagnostic procedures. Prior to the outage, nearly all the Mo99 produced for the world market came from five aging government owned research reactors in Canada, France, the Netherlands, Belgium and South Africa. The NRU, the largest of these, produced about 30 to 40 percent of the world supply of isotopes prior to 2009 - since its return to service in 2010, its world market share is estimated at 15 to 20%.

  18. Two isotopic methods for estimation of soybean nitrogen fixation

    International Nuclear Information System (INIS)

    Domenach, A.-M.; Chalamet, Alain; Pachiaudi, Christiane

    1979-01-01

    Measurements of rate of nitrogen symbiotic fixation by Soybean were determined by two different methods: variations in the natural abundance of 15 N in plants; use of 'A' value determined from labelled nitrogen applications to the soil. The results from both methods were in good agreement. Rates of fixed nitrogen were similar when using non nodulated Soybean or Ray-Grass as reference [fr

  19. Two isotopic methods for estimation of soybean nitrogen fixation

    Energy Technology Data Exchange (ETDEWEB)

    Domenach, A M; Chalamet, A; Pachiaudi, C [Lyon-1 Univ., 69 - Villeurbanne (France)

    1979-07-16

    Measurements of rate of nitrogen symbiotic fixation by Soybean were determined by two different methods: variations in the natural abundance of /sup 15/N in plants; use of 'A' value determined from labelled nitrogen applications to the soil. The results from both methods were in good agreement. Rates of fixed nitrogen were similar when using non nodulated Soybean or Ray-Grass as reference.

  20. Benzylic rearrangement stable isotope labeling for quantitation of guanidino and ureido compounds in thyroid tissues by liquid chromatography-electrospray ionization mass spectrometry

    International Nuclear Information System (INIS)

    Fan, Ruo-Jing; Guan, Qing; Zhang, Fang; Leng, Jia-Peng; Sun, Tuan-Qi; Guo, Yin-Long

    2016-01-01

    Benzylic rearrangement stable isotope labeling (BRSIL) was explored to quantify the guanidino and ureido compounds (GCs and UCs). This method employed a common reagent, benzil, to label the guanidino and ureido groups through nucleophilic attacking then benzylic migrating. The use of BRSIL was investigated in the analysis of five GCs (creatine, L-arginine, homoarginine, 4-guanidinobutyric acid, and methylguanidine) and two UCs (urea and citrulline). The labeling was found simple and specific. The introduction of bi-phenyl group and the generation of nitrogen heterocyclic ring in the benzil-d0/d5 labeled GCs and UCs improved the retention behaviors in liquid chromatography (LC) and increased the sensitivity of electrospray ionization mass spectrometry (ESI MS) detection. The fragment ion pairs of m/z 182/187 and m/z 210/215 from the benzil-d0/d5 tags facilitated the discovery of potential GCs and UCs candidates residing in biological matrices. The use of BRSIL combined with LC-ESI MS was applied for simultaneously quantitation of GCs and UCs in thyroid tissues. It was demonstrated that nine GCs and UCs were detected, six of which were further quantified based on corresponding standards. It was concluded that five GCs and UCs (L-arginine, homoarginine, 4-guanidinobutyric acid, methylguanidine, and citrulline) were statistically significantly different (p < 0.05) between the para-carcinoma and carcinoma thyroid tissue samples. - Highlights: • A common reagent, benzil-d0/d5 was employed to label the GCs and UCs through BRSIL. • The benzil-d0/d5 labeling improved the retention behavior in RPLC and increased the sensitivity by ESI MS detection. • BRSIL coupled with LC-ESI MS was applied to the qualitation and quantitation of GCs and UCs in thyroid tissues.

  1. Benzylic rearrangement stable isotope labeling for quantitation of guanidino and ureido compounds in thyroid tissues by liquid chromatography-electrospray ionization mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Fan, Ruo-Jing [State Key Laboratory of Organmetallic Chemistry and National Center for Organic Mass Spectrometry in Shanghai, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, 345 Lingling Road, Shanghai, 200032 (China); Guan, Qing [Department of Head and Neck Surgery, Fudan University Shanghai Cancer Center, Shanghai, 200032 (China); Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032 (China); Zhang, Fang, E-mail: fzhang@sioc.ac.cn [State Key Laboratory of Organmetallic Chemistry and National Center for Organic Mass Spectrometry in Shanghai, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, 345 Lingling Road, Shanghai, 200032 (China); Leng, Jia-Peng [State Key Laboratory of Organmetallic Chemistry and National Center for Organic Mass Spectrometry in Shanghai, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, 345 Lingling Road, Shanghai, 200032 (China); Sun, Tuan-Qi, E-mail: tuanqisun@163.com [Department of Head and Neck Surgery, Fudan University Shanghai Cancer Center, Shanghai, 200032 (China); Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032 (China); Guo, Yin-Long, E-mail: ylguo@sioc.ac.cn [State Key Laboratory of Organmetallic Chemistry and National Center for Organic Mass Spectrometry in Shanghai, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, 345 Lingling Road, Shanghai, 200032 (China)

    2016-02-18

    Benzylic rearrangement stable isotope labeling (BRSIL) was explored to quantify the guanidino and ureido compounds (GCs and UCs). This method employed a common reagent, benzil, to label the guanidino and ureido groups through nucleophilic attacking then benzylic migrating. The use of BRSIL was investigated in the analysis of five GCs (creatine, L-arginine, homoarginine, 4-guanidinobutyric acid, and methylguanidine) and two UCs (urea and citrulline). The labeling was found simple and specific. The introduction of bi-phenyl group and the generation of nitrogen heterocyclic ring in the benzil-d0/d5 labeled GCs and UCs improved the retention behaviors in liquid chromatography (LC) and increased the sensitivity of electrospray ionization mass spectrometry (ESI MS) detection. The fragment ion pairs of m/z 182/187 and m/z 210/215 from the benzil-d0/d5 tags facilitated the discovery of potential GCs and UCs candidates residing in biological matrices. The use of BRSIL combined with LC-ESI MS was applied for simultaneously quantitation of GCs and UCs in thyroid tissues. It was demonstrated that nine GCs and UCs were detected, six of which were further quantified based on corresponding standards. It was concluded that five GCs and UCs (L-arginine, homoarginine, 4-guanidinobutyric acid, methylguanidine, and citrulline) were statistically significantly different (p < 0.05) between the para-carcinoma and carcinoma thyroid tissue samples. - Highlights: • A common reagent, benzil-d0/d5 was employed to label the GCs and UCs through BRSIL. • The benzil-d0/d5 labeling improved the retention behavior in RPLC and increased the sensitivity by ESI MS detection. • BRSIL coupled with LC-ESI MS was applied to the qualitation and quantitation of GCs and UCs in thyroid tissues.

  2. Raman scattering method and apparatus for measuring isotope ratios and isotopic abundances

    International Nuclear Information System (INIS)

    Harney, R.C.; Bloom, S.D.

    1978-01-01

    Raman scattering is used to measure isotope ratios and/or isotopic abundances. A beam of quasi-monochromatic photons is directed onto the sample to be analyzed, and the resulting Raman-scattered photons are detected and counted for each isotopic species of interest. These photon counts are treated mathematically to yield the desired isotope ratios or isotopic abundances

  3. Stable isotope N-phosphoryl amino acids labeling for quantitative profiling of amine-containing metabolites using liquid chromatography mass spectrometry.

    Science.gov (United States)

    Zhang, Shanshan; Shi, Jinwen; Shan, Changkai; Huang, Chengting; Wu, Yile; Ding, Rong; Xue, Yuhua; Liu, Wen; Zhou, Qiang; Zhao, Yufen; Xu, Pengxiang; Gao, Xiang

    2017-07-25

    Stable isotope chemical labeling liquid chromatography-mass spectrometry (LC-MS) is a powerful strategy for comprehensive metabolomics profiling, which can improve metabolites coverage and quantitative information for exploration of metabolic regulation in complex biological systems. In the current work, a novel stable isotope N-phosphoryl amino acids labeling strategy (SIPAL) has been successful developed for quantitative profiling of amine-containing metabolites in urine based on organic phosphorus chemistry. Two isotopic reagents, 16 O 2 - and 18 O 2 -N-diisopropyl phosphoryl l-alanine N-hydroxysuccinimide esters ( 16 O/ 18 O-DIPP-L-Ala-NHS), were firstly synthesized in high yields for labeling the amine-containing metabolites. The performance of SIPAL strategy was tested by analyzing standard samples including 20 l-amino acids, 10 d-amino acids and small peptides by using LC-MS. We observed highly efficient and selective labeling for SIPAL strategy within 15 min in a one-pot derivatization reaction under aqueous reaction conditions. The introduction of a neutral phosphate group at N-terminus can increase the proton affinity and overall hydrophobicity of targeted metabolites, leading to the better ionization efficiency in electrospray ionization processes and chromatographic separations of hydrophilic metabolites on reversed-phase column. Furthermore, the chiral metabolites, such as d-amino acids, could be converted to diastereomers after SIPAL and successfully separated on regular reversed-phase column. The chirality of labeled enantiomers can be determined by using different detection methods such as 31 P NMR, UV, and MS, demonstrating the potential application of SIPAL strategy. In addition, absolute quantification of chiral metabolites in biological samples can be easily achieved by using SIPAL strategy. For this purpose, urine samples collected from a healthy volunteer were analyzed by using LC-ESI-Orbitrap MS. Over 300 pairs of different amine

  4. Isotopic labeling as a tool to establish intramolecular vibrational coupling: The reaction of 2-propanol on Mo(110)

    International Nuclear Information System (INIS)

    Uvdal, P.; Wiegand, B.C.; Serafin, J.G.; Friend, C.M.

    1992-01-01

    The reactions of 2-propanol on Mo(110) were investigated using temperature programmed reaction, high resolution electron energy loss, and x-ray photoelectron spectroscopies. 2-Propanol forms 2-propoxide upon adsorption at 120 K on Mo(110). The 2-propoxide intermediate deoxygenates via selective γ C--H bond scission to eliminate propene as well as C--O bond hydrogenolysis to form trace amounts of propane. The C--O bond of 2-propoxide is estimated to be nearly perpendicular to the surface. Selective isotopic labeling was used to establish the coupling between the C--O stretch and modes associated with the hydrocarbon framework. The degree of coupling was strongly affected by bonding to the surface, primarily due to weakening of the C--O bond when 2-propoxide is bound to Mo(110). Selective isotopic labeling was, therefore, essential in making vibrational assignments and in identifying key reaction steps. Only a small kinetic isotope effect was observed during reaction of (CD 3 )(CH 3 )CHOH, consistent with a substantial component of C--O bond breaking in the transition state for propene elimination. Coupling of the C--O stretch to motion of the methyl group is also suggested to be important in the transition state for propene elimination

  5. Community Mining Method of Label Propagation Based on Dense Pairs

    Directory of Open Access Journals (Sweden)

    WENG Wei

    2014-03-01

    Full Text Available In recent years, with the popularity of handheld Internet equipments like mobile phones, increasing numbers of people are becoming involved in the virtual social network. Because of its large amount of data and complex structure, the network faces new challenges of community mining. A label propagation algorithm with low time complexity and without prior parameters deals easily with a large networks. This study explored a new method of community mining, based on label propagation with two stages. The first stage involved identifying closely linked nodes according to their local adjacency relations that gave rise to a micro-community. The second stage involved expanding and adjusting this community through a label propagation algorithm (LPA to finally obtain the community structure of the entire social network. This algorithm reduced the number of initial labels and avoided the merging of small communities in general LPAs. Thus, the quality of community discovery was improved, and the linear time complexity of the LPA was maintained.

  6. Development of radio-labeling method for natural juvenile hormone

    International Nuclear Information System (INIS)

    Kurata, Keiji; Shiozuki, Takahiro; Kotaki, Toyomi

    1997-01-01

    The aim of this study was to develop a new method for quantitative determination of juvenile hormone (JH) based on the principle for radioimmuno assay. Using JH-binding protein (JHBP), the discrimination of the L-form of JH from D-form not occurring naturally was attempted to establish a measuring method for JH. First, the corpus allatum, the JH-producing endocrine organ was cultured and both L-form JH and 3 H or 14 C-labelled JH could be obtained easily. There are several homologs of JH and it is necessary to establish the respective labelling methods for the homologues. Since different species of insects produce JH with its specific structure, each homologue could be produced by selecting an appropriate species. The capacity of JH production was compared among five insects. The biosynthetic ability by the corpus allatum from migratory locust was highest among them and 1 μg of labelled JH type 3 could be obtained from the culture with about 50 corpus allata for 3 hrs. Since other materials than JH were also released into the culture medium, establishment of the effective method for isolation and purification of JH is necessary to use it for bioassay. (M.N.)

  7. Analysis of the differentially expressed low molecular weight peptides in human serum via an N-terminal isotope labeling technique combining nano-liquid chromatography/matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Leng, Jiapeng; Zhu, Dong; Wu, Duojiao; Zhu, Tongyu; Zhao, Ningwei; Guo, Yinlong

    2012-11-15

    Peptidomics analysis of human serum is challenging due to the low abundance of serum peptides and interference from the complex matrix. This study analyzed the differentially expressed (DE) low molecular weight peptides in human serum integrating a DMPITC-based N-terminal isotope labeling technique with nano-liquid chromatography and matrix-assisted laser desorption/ionization mass spectrometry (nano-LC/MALDI-MS). The workflow introduced a [d(6)]-4,6-dimethoxypyrimidine-2-isothiocyanate (DMPITC)-labeled mixture of aliquots from test samples as the internal standard. The spiked [d(0)]-DMPITC-labeled samples were separated by nano-LC then spotted on the MALDI target. Both quantitative and qualitative studies for serum peptides were achieved based on the isotope-labeled peaks. The DMPITC labeling technique combined with nano-LC/MALDI-MS not only minimized the errors in peptide quantitation, but also allowed convenient recognition of the labeled peptides due to the 6 Da mass difference. The data showed that the entire research procedure as well as the subsequent data analysis method were effective, reproducible, and sensitive for the analysis of DE serum peptides. This study successfully established a research model for DE serum peptides using DMPITC-based N-terminal isotope labeling and nano-LC/MALDI-MS. Application of the DMPITC-based N-terminal labeling technique is expected to provide a promising tool for the investigation of peptides in vivo, especially for the analysis of DE peptides under different biological conditions. Copyright © 2012 John Wiley & Sons, Ltd.

  8. Absorption of food cobalamins assessed by the double isotope method in healthy volunteers and in patients with chronic diarrhoea

    International Nuclear Information System (INIS)

    Kittang, B.; Hamborg, B.; Schjoensby, H.

    1985-01-01

    To make a food preparation containing radioactively labelled cobalamins, rabbits were given repeated injections with 57 Co-labelled cyanocobalamin. The liver was removed, homogenized and fried for one min. or boiled for 30 min. Of the radioactivity in the fried homogenate, 41.7% was recovered in the centrifuged supernatant compared with 50.8% in the boiled homogenate. The radioactivity in the supernatants had a molecular size close to that of free 57 Co-labelled cyanocobalamin. 42% of the radioactivity in the whole homogenate had been incorporated into 5-deoxyadenosyl-, 10% into methyl-and 16.5% into hydroxy-cobalamin. To assess the validity of a double-isotope method for measuring the intestinal absorption of doses of 57 Co-labelled liver cobalamins, 51 CrCl 3 was used as a non-absorbable marker. In 14 healthy volunteers the correlation coefficient between the absorption measured by the double-isotope technique and the faecal excretion test was highly significant (r = 0.96, p 57 Co/ 51 Cr ratio in successive stool collections. In 11 patients with chronic diarrhoea there was a significant correlation between the absorption measured by the double-isotope technique and the faecal excretion test (r = 0.92, p 57 / 51 Cr ratio in successive stool collections

  9. Bacterial diversity exploration in hydrocarbon polluted soil: metabolic potential and degrader community evolution revealed by isotope labeling

    International Nuclear Information System (INIS)

    Martin, F.

    2011-01-01

    Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous compounds produced by incomplete combustion of organic matter. They are a source of environmental pollution, especially associated to oil product exploitation, and represent a threat for living organisms including human beings because of their toxicity. Many bacteria capable of degrading PAHs have been isolated and studied. However, since less than 5% of soil bacteria can be cultivated in the laboratory, bacterial species able to degrade PAHs in situ have been poorly studied. The first goal of this study was to identify bacteria that degrade PAHs in soil using culture-independent molecular methods. To this end, a strategy known a stable isotope probing has been implemented based on the use of phenanthrene, a three rings PAH, in which the natural isotope of carbon was replaced by 13 C. This molecule has been introduced as a tracer in microcosms containing soil from a constructed wetlands collecting contaminated water from highway runoff. Bacteria having incorporated the 13 C were then identified by 16 S rRNA gene sequence analysis after PCR amplification from labeled genomic DNA extracted from soil. The results show that so far little studied Betaproteobacteria, belonging to the genera Acidovorax, Rhodoferax, Hydrogenophaga and Thiobacillus, as well as Rhodocyclaceae, were the key players in phenanthrene degradation. Predominance of Betaproteobacteries was established thanks to quantitative PCR measurements. A dynamic analysis of bacterial diversity also showed that the community structure of degraders depended on phenanthrene bioavailability. In addition, the phylogenetic diversity of ring-hydroxylating di-oxygenases, enzymes involved in the first step of PAH degradation, has been explored. We detected new sequences, mostly related to di-oxygenases from Sphingomonadales and Burkholderiales. For the first time, we were able to associate a catalytic activity for oxidation of PAHs to partial gene sequences

  10. Method of deuterium isotope separation and enrichment

    International Nuclear Information System (INIS)

    Benson, S.W.

    1978-01-01

    A method of separating deuterium, i.e., heavy hydrogen, from certain naturally occurring sources using tuned infrared lasers to selectively decompose specified classes of organic molecules (i.e., RX) into enriched molecular products containing deuterium atoms is described. The deuterium containing molecules are easily separated from the starting material by absorption, distillation or other simple chemical separation techniques and methods. After evaporation such deuterium containing molecules can be burned to form water with an enriched deuterium content or pyrolyzed to form hydrogen gas with an enriched deuterium content. The undecomposed molecules and the other reaction products which are depleted of their deuterium containing species can be catalytically treated, preferably using normal water, to restore the natural abundance of deuterium and such restored molecules can then be recycled

  11. Synthesis of isotopically labelled angiotensin II receptor antagonist GR138950X

    International Nuclear Information System (INIS)

    Carr, R.M.; Cable, K.M.; Newman, J.J.; Sutherland, D.R.

    1996-01-01

    Syntheses of [ 13 C] and [ 14 C]-labelled versions of angiotensin II receptor antagonist GR138950X, labelled in the imidazole carboxamide residue, are described. These involved preparation of an iodoimidazole substrate by a novel iododecarboxylation procedure, followed by cyanation with a mixture of carbon-labelled potassium cyanide and copper (l) iodide in DMF at high temperature. The preparation of a mass-labelled (M+5) version of GR138950X is also described. This involved the synthesis of an [ 13 C 3 , 15 N 2 ]-labelled imidazole from a 1,2,3-tricarbonyl compound, [ 13 C 3 ]propionaldehyde and [ 15 N]ammonia. The labelled imidazole was further elaborated into multiply-labelled GR138950X. (Author)

  12. Optimizing design parameter for light isotopes separation by distillation method

    International Nuclear Information System (INIS)

    Ahmadi, M.

    1999-01-01

    More than methods are suggested in the world for producing heavy water, where between them chemical isotopic methods, distillation and electro lys are used widely in industrial scale. To select suitable method for heavy water production in Iran, taking into consideration, domestic technology an facilities, combination of hydrogen sulphide-water dual temperature process (Gs) and distillation (D W) may be proposed. Natural water, is firstly enriched up to 15 a% by G S process and then by distillation unit is enriched up to the grade necessary for Candu type reactors (99.8 a%). The aim of present thesis, is to achieve know-how, optimization of design parameters, and executing basic design for water isotopes separation using distillation process in a plant having minimum scale possible. In distillation, vapour phase resulted from liquid phase heating, is evidently composed of the same constituents as liquid phase. In isotopic distillation, the difference in composition of constituents is not considerable. In fact alteration of constituents composition is so small that makes the separation process impossible, however, direct separation and production of pure products without further processing which becomes possible by distillation, makes this process as one of the most important separation processes. Profiting distillation process to produce heavy water is based on difference existing between boiling point of heavy and light water. The trends of boiling points differences (heavy and light water) is adversely dependant with pressure. As the whole system pressure decreases, difference in boiling points increases. On the other hand according to the definition, separation factor is equal to the ratio of pure light water vapour pressure to that of heavy water, or we can say that the trend of whole system pressure decrease results in separation factor increase, which accordingly separation factor equation to pressure variable should be computed firstly. According to the

  13. Overcoming interference with the detection of a stable isotopically labeled microtracer in the evaluation of beclabuvir absolute bioavailability using a concomitant microtracer approach.

    Science.gov (United States)

    Jiang, Hao; Titsch, Craig; Zeng, Jianing; Jones, Barry; Joyce, Philip; Gandhi, Yash; Turley, Wesley; Burrell, Richard; Aubry, Anne F; Arnold, Mark E

    2017-09-05

    The oral absolute bioavailability of beclabuvir in healthy subjects was determined using a microdose (100μg) of the stable isotopically labeled tracer via intravenous (IV) infusion started after oral dosing of beclabuvir (150mg). To simultaneously analyze the concentrations of the IV microtracer ([ 13 C 6 ]beclabuvir) and beclabuvir in plasma samples, a liquid chromatography-triple quadrupole mass spectrometry (LC-MS/MS) method was initially developed. Surprisingly beclabuvir significantly interfered with the IV microtracer detection when using the selected reaction monitoring (SRM) in the assay. An interfering component from the drug substance was observed using a high resolution mass spectrometer (HRMS). The mass-to-charge (m/z) of the interfering component was -32ppm different from the nominal value for the IV microtracer and thus could not be differentiated in the SRM assay by the unit mass resolution. To overcome this interference, we evaluated two approaches by either monitoring an alternative product ion using the SRM assay or isolating the interfering component using the parallel reaction monitoring (PRM) assay on the HRMS. This case study has demonstrated two practical approaches for overcoming interferences with the detection of stable isotopically labeled IV microtracers in the evaluation of absolute bioavailability, which provides users the flexibility in using either LC-MS/MS or HRMS to mitigate unpredicted interferences in the assay to support microtracer absolute bioavailability studies. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Method of deuterium isotope separation and enrichment

    International Nuclear Information System (INIS)

    Benson, S.W.

    1979-01-01

    A method is described for separating and enriching deuterium containing molecules comprising the steps of: providing a source of organic molecules containing a normal abundance of deuterium atoms, the organic molecules having a structural formula RX, in which R is an organic radical selected from ethyl, isopropyl, t-butyl and 3-cyclopentenyl, and in which X is selected from F, Cl, Br and OH, and wherein R represents 3-cyclopentenyl, X may additionally represent H; exposing the molecules to the radiation of at least one pulsed infrared laser source which has been specifically tuned and focussed to selectively decompose RX molecules containing deuterium to form an enriched olefin specie containing deuterium, and HX; and separating the deuterium enriched olefin specie from the undecomposed deuterium depleted RX molecules and HX. (author)

  15. Advisory group meeting on stable isotope labelled compounds in biomedical studies

    International Nuclear Information System (INIS)

    Vera Ruiz, H.; Parr, R.M.

    1985-11-01

    The programme of the meeting was restricted to topics involving applications of stable isotopes of the lighter elements (H, C, N, O). The current status of stable isotope techniques and applications in nutritional and biomedical studies, the applicability of these techniques in developing countries and the IAEA's future programmes on this topic were discussed

  16. A method of computerised isotope dynamic proctography

    International Nuclear Information System (INIS)

    Papachrysostomou, M.; Ferrington, C.; Merrick, M.V.; Smith, A.N.; Western General Hospital, Edinburgh; Griffin, T.M.J.

    1992-01-01

    Patients with long-standing constipation were examined by radioisotope proctography. A radiolabelled synthetic potato mash was inserted intrarectally and the dynamic changes during simulated defaecation recorded using a gamma-camera. Computer images from the stored data illustrate changes in the anorectal angle and pelvic floor. The median (and 95% confidence intervals) of the anorectal angles were: At rest 105deg (101deg, 116deg), on voluntary contraction of the pelvic floor by 'squeezing' 91deg (81deg, 98deg), on straining 120deg (107deg, 137deg) and during evacuation 126deg (116deg, 153deg). The pelvic floor movements were: Pelvic floor ascent on voluntary contraction 28 mm (9, 34 mm), pelvic floor descent on straining -8 mm (-14, -4 mm) and descent during evacuation -27 mm (-34, -11 mm). Useful additions to previous methods are measurement of the completeness of rectal evacuation 58% (42, 77 mm), the defaecation time 64 s (50, 138 s) and the defaecation rate 0.9%/s (0.4, 1.4%/s). (orig.)

  17. Analytical method for the isotopic characterization of soils

    International Nuclear Information System (INIS)

    Sibello Hernandez, Rita; Cozzella, Maria Letizia; Mariani, Mario

    2014-01-01

    The aim of this work was to develop an analytical method in order to determine the isotopic composition of different elements in soil samples and to determine the existence of contamination. The method used in the digestion of the samples was the EPA 3050B, and some metal concentration were determined including uranium and thorium. For elements with even lower concentrations such as plutonium and radium a treatment after mineralization by EPA, was necessary. The measurement technique used was mass spectrometry with quadrupole and plasma induced associated (ICP-MS). Results of the analysis performed in two laboratories showed a good correspondence. This method allowed to perform the isotopic characterization of studied soils and results showed that the studied soils do not present any local pollution and that the presence of plutonium-239, is due to global failure

  18. Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC) Applied to Quantitative Proteomics of Bacillus subtilis

    DEFF Research Database (Denmark)

    Soufi, Boumediene; Kumar, C.; Gnad, F.

    2010-01-01

    We applied stable isotope labeling by amino acids in cell culture (SILAC) to large-scale quantitative proteomics analyses of the model bacterium Bacillus subtilis in two physiological conditions: growth on succinate and growth under phosphate starvation. Using a B. subtilis strain auxotrophic...... of the most comprehensive quantitative proteomics studies in bacteria, covering more than 75% of the B. subtilis genes expressed in the log phase of growth. Furthermore, we detect and quantify dynamics of 35 Ser/Thr/Tyr phosphorylation sites under growth on succinate, and 10 phosphorylation sites under...

  19. A Cost-effective Amino-acid-type Selective Isotope Labeling of Proteins Expressed in Leishmania tarentolae

    Czech Academy of Sciences Publication Activity Database

    Foldynová-Trantírková, Silvie; Matulová, J.; Dötsch, V.; Löhr, F.; Cirstea, I.; Alexandov, K.; Breitling, R.; Lukeš, Julius; Trantírek, Lukáš

    2009-01-01

    Roč. 26, č. 6 (2009), s. 755-761 ISSN 0739-1102 R&D Projects: GA ČR GP204/08/P585; GA AV ČR 1QS600220554; GA AV ČR KAN200100801; GA MŠk 2B06129 Institutional research plan: CEZ:AV0Z60220518 Keywords : NMR * isotope labeling * protein expression * Leishmania * low-level enrichment Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.124, year: 2009

  20. Evaluation of a new component used for isotopic lymphography: colloidal rhenium sulfide sup(99m)Tc labelled

    International Nuclear Information System (INIS)

    Pecking, A.; Le Mercier, N.; Gobin, R.; Bardy, A.; Najean, Y.

    1978-01-01

    We have studied for lymphatic scintigraphy a new radiopharmaceutical, sup(99m)Tc-labelled rhenium sulfocolloid. This preliminary study includes 20 adults patients with lymphomas and lymphoedemas. The principal advantage of this drug is its absence of toxicity and local pain, so that a rapid sub-cutaneous injection without local anesthesia is made possible. Good results have been obtained, as well in morphological studies of para-aortic and mammary lymph nodes as for kinetic studies of lymphatic flow in lymphoedemas. No liver and spleen uptake of radio-isotope was observed after foot injection [fr

  1. Simultaneous determination of the intravenous and oral pharmacokinetic parameters of D,L-verapamil using stable isotope-labelled verapamil.

    Science.gov (United States)

    Eichelbaum, M; Somogyi, A; von Unruh, G E; Dengler, H J

    1981-01-01

    Following i.v. administration, the plasma concentration-time curve of verapamil could best be described by either a mono- or biexponential equation. Total plasma clearance (1.26 1/min) approached liver blood flow (1.51/min), so it can be concluded that its clearance is liver blood flow-dependent. Although absorption was almost complete after oral administration, absolute bioavailability (20%) was low, due to extensive hepatic first-pass metabolism. The approach using stable isotope-labelled and unlabelled drug permits simultaneous administration by the intravascular and extravascular routes, thus allowing determination of absolute bioavailability in a single experiment.

  2. A new method for studying iodine metabolism; the isotopic equilibrium method - kinetic and quantitative aspects of measurements made on rats

    International Nuclear Information System (INIS)

    Simon, C.

    1964-05-01

    The isotopic equilibrium method which has been developed in the case of the rat has made it possible to measure the absolute values of the principal parameters of iodine metabolism in this animal. The quantities and concentrations of iodine have been measured in the thyroid gland and in the plasma with a sensitivity of 0.001 μg of 127 I. This sensitivity has made it possible to measure pools as small as the iodide and the free iodotyrosines of the thyroid and to demonstrate the absence of free iodotyrosines in the plasma of the normal rat. In vivo, the isotopic equilibrium method has made it possible to measure the iodine content of the thyroid gland and to calculate the intensity of this gland's secretion without removing it. By double labelling with 125 I and 131 I the isotopic equilibrium method has made it possible to measure the flux, intensity of the intrathyroidal recycling as well as the turnover rates of all the iodine containing compounds of the thyroid gland. For this gland no precursor-product relationship has been found between The iodotyrosines (MIT and DIT) and the iodothyronines (T 4 and T 3 ). The absence of this relationship is due to the heterogeneity of the thyroglobulin turnover. It has been shown furthermore that there exists in the plasma an organic fraction of the iodine which is different to thyroglobulin and which is renewed more rapidly than the circulating hormones T 3 and T 4 . The isotopic equilibrium method is very useful for series measurements of iodine. It makes it possible furthermore to improve the biochemical fractionations by adding carriers without affecting the subsequent 127 I measurements. (author) [fr

  3. Isotope correlations for safeguards surveillance and accountancy methods

    International Nuclear Information System (INIS)

    Persiani, P.J.; Kalimullah.

    1983-01-01

    Isotope correlations corroborated by experiments, coupled with measurement methods for nuclear material in the fuel cycle have the potential as a safeguards surveillance and accountancy system. The US/DOE/OSS Isotope Correlations for Surveillance and Accountancy Methods (ICSAM) program has been structured into three phases: (1) the analytical development of Isotope Correlation Technique (ICT) for actual power reactor fuel cycles; (2) the development of a dedicated portable ICT computer system for in-field implementation, and (3) the experimental program for measurement of U, Pu isotopics in representative spent fuel-rods of the initial 3 or 4 burnup cycles of the Commonwealth Edison Zion -1 and -2 PWR power plants. Since any particular correlation could generate different curves depending upon the type and positioning of the fuel assembly, a 3-D reactor model and 2-group cross section depletion calculation for the first cycle of the ZION-2 was performed with each fuel assembly as a depletion block. It is found that for a given PWR all assemblies with a unique combination of enrichment zone and number of burnable poison rods (BPRs) generate one coincident curve. Some correlations are found to generate a single curve for assemblies of all enrichments and number of BPRs. The 8 axial segments of the 3-D calculation generate one coincident curve for each correlation. For some correlations the curve for the full assembly homogenized over core-height deviates from the curve for the 8 axial segments, and for other correlations coincides with the curve for the segments. The former behavior is primarily based on the transmutation lag between the end segment and the middle segments. The experimental implication is that the isotope correlations exhibiting this behavior can be determined by dissolving a full assembly but not by dissolving only an axial segment, or pellets

  4. Metabolic cartography: experimental quantification of metabolic fluxes from isotopic labelling studies.

    Science.gov (United States)

    O'Grady, John; Schwender, Jörg; Shachar-Hill, Yair; Morgan, John A

    2012-03-01

    For the past decade, flux maps have provided researchers with an in-depth perspective on plant metabolism. As a rapidly developing field, significant headway has been made recently in computation, experimentation, and overall understanding of metabolic flux analysis. These advances are particularly applicable to the study of plant metabolism. New dynamic computational methods such as non-stationary metabolic flux analysis are finding their place in the toolbox of metabolic engineering, allowing more organisms to be studied and decreasing the time necessary for experimentation, thereby opening new avenues by which to explore the vast diversity of plant metabolism. Also, improved methods of metabolite detection and measurement have been developed, enabling increasingly greater resolution of flux measurements and the analysis of a greater number of the multitude of plant metabolic pathways. Methods to deconvolute organelle-specific metabolism are employed with increasing effectiveness, elucidating the compartmental specificity inherent in plant metabolism. Advances in metabolite measurements have also enabled new types of experiments, such as the calculation of metabolic fluxes based on (13)CO(2) dynamic labelling data, and will continue to direct plant metabolic engineering. Newly calculated metabolic flux maps reveal surprising and useful information about plant metabolism, guiding future genetic engineering of crops to higher yields. Due to the significant level of complexity in plants, these methods in combination with other systems biology measurements are necessary to guide plant metabolic engineering in the future.

  5. Enantio-specific C(sp3)-H activation catalyzed by ruthenium nanoparticles: application to isotopic labeling of molecules of biological interest

    International Nuclear Information System (INIS)

    Taglang, Celine

    2015-01-01

    Isotopic labeling with deuterium and tritium is extensively used in chemistry, biology and pharmaceutical research. Numerous methods of labeling by isotopic exchange allow high isotopic enrichments but generally require harsh conditions (high temperatures, acidity). As a consequence, a general, regioselective and smooth labeling method that might be applicable to a wide diversity of substrates remains to develop. In the first part of this thesis, we demonstrated that the use of ruthenium nanoparticles, synthesized by Pr. Bruno Chaudret's team (INSA Toulouse), allowed the mild (2 bar of deuterium gas at 55 C), effective and selective H/D exchange reaction of a large variety of nitrogen-containing compounds, such as pyridines, indoles and primary, secondary and tertiary alkyl amines. The usefulness and the efficiency of this novel methodology was demonstrated by the deuteration of eight nitrogen-containing molecules of biological interest without altering their chemical and stereochemical properties. However, the conservation of the original stereochemistry of an activated chiral C-H center remains a major issue. We studied the reactivity of RuNP(at)PVP on different categories of nitrogen-containing substrates (amines, aminoacids and peptides) in water or in organic solvents. Our results showed that C-H activation of chiral carbons C(sp3) took place efficiently, selectively and, in all cases, with total retention of configuration. The wide range of applications of this procedure was demonstrated by the labeling of three chiral amines, fourteen aminoacids, three aromatic amino esters and four peptides. Moreover, our collaboration with Pr. Romuald Poteau's team (INSA Toulouse) led to the identification of two mechanisms by ab initio simulation in agreement with our experimental results: the σ-bond metathesis mechanism and the oxidative addition mechanism. These two mechanisms imply two vicinal ruthenium atoms leading to the formation an original

  6. METHOD FOR PRODUCING ISOTOPIC METHANES AND PARTIALLY HALOGENATED DERIVATIVES THEROF

    Science.gov (United States)

    Frazer, J.W.

    1959-08-18

    A method is given for producing isotopic methanes and/ or partially halogenated derivatives. Lithium hydride, deuteride, or tritide is reacted with a halogenated methane or with a halogenated methane in combination with free halogen. The process is conveniently carried out by passing a halogenated methane preferably at low pressures or in an admixture with an inert gas through a fixed bed of finely divided lithium hydride heated initially to temperatures of 100 to 200 deg C depending upon the halogenated methane used.

  7. Isotopic method of leaks detection in oil pipelines

    International Nuclear Information System (INIS)

    Listwam, W.; Mottl, J.

    1974-01-01

    Isotopic method of leaks detection in oil pipelines of diameter 200-800 mm is described. Tracer is injected into pipeline in the form of CH 3 Br 82 . After few hours one or two detectors are passed through pipeline to detect leaks. Detector set consists of scintillation radiometer with Na I/Tl crystal, electronic blecks with one-channel analyzer, recorder and storage batteries. Detector set is built on integrated circuits. (Z.M.)

  8. Direct infusion electrospray ionization–ion mobility–mass spectrometry for comparative profiling of fatty acids based on stable isotope labeling

    Energy Technology Data Exchange (ETDEWEB)

    Leng, Jiapeng, E-mail: jpleng@126.com [Department of Chemical Engineering, North University of China, Taiyuan 030051 (China); Guan, Qing [Department of Head and Neck Surgery, Fudan University Shanghai Cancer Center (FUSCC), Shanghai 200032 (China); Department of Oncology, Shanghai Medical College, Fudan University, Shanghai 200032 (China); Sun, Tuanqi, E-mail: tuanqisun@163.com [Department of Head and Neck Surgery, Fudan University Shanghai Cancer Center (FUSCC), Shanghai 200032 (China); Department of Oncology, Shanghai Medical College, Fudan University, Shanghai 200032 (China); Wang, Haoyang [National Center for Organic Mass Spectrometry in Shanghai, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai 200032 (China); Cui, Jianlan; Liu, Qinghao [Department of Chemical Engineering, North University of China, Taiyuan 030051 (China); Zhang, Zhixu; Zhang, Manyu [Agilent Technologies China Co., Ltd, Shanghai 200080 (China); Guo, Yinlong, E-mail: ylguo@sioc.ac.cn [National Center for Organic Mass Spectrometry in Shanghai, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai 200032 (China)

    2015-08-05

    A rapid method for fatty acids (FAs) comparative profiling based on carboxyl-specific stable isotope labeling (SIL) and direct infusion electrospray ionization–ion mobility–mass spectrometry (ESI–IM–MS) is established. The design of the method takes advantage of the three-dimensional characteristics of IM–MS including drift time, m/z and ion intensity, for comparison of d0-/d6-2,4-dimethoxy-6-piperazin-1-yl pyrimidine (DMPP)-labeled FAs. In particular, without chromatographic separation, the method allowed direct FAs profiling in complex samples due to the advantageous priority of DMPP in signal enhancement as well as the extra resolution that IM–MS offered. Additionally, the d0-/d6-DMPP-labeled FAs showed expected features, including very similar drift times, 6 Da mass deviations, specific reporter ions, similar MS responses, and adherence to the drift time rule regarding the influence of carbon chain length and unsaturation on relative drift times. Therefore, the introduction of isotope analogs minimized the matrix effect and variations in quantification and ensured accurate identification of non-targeted FAs by those typical features. Peak intensity ratios between d0-/d6-DMPP-labeled ions were subsequently used in relative quantification for the detected FAs. The established strategy has been applied successfully in the rapid profiling of trace free FAs between normal and cancerous human thyroid tissues. Sixteen free FAs were found with the increased level with a statistically significant difference (p < 0.05) compared to the normal tissue samples. The integrated SIL technique and ESI–IM–MS are expected to serve as an alternative tool for high-throughput analysis of FAs in complex samples. - Highlights: • A novel method based on IM–MS and SIL was developed for FAs comparative profiling. • Without LC separation, the method allowed direct infusion profiling of FAs in complex samples. • Both of the efficiency and accuracy for FAs analyses

  9. Isotope correlations for safeguards surveillance and accountancy methods

    International Nuclear Information System (INIS)

    Persiani, P.J.; Kalimullah.

    1982-01-01

    Isotope correlations corroborated by experiments, coupled with measurement methods for nuclear material in the fuel cycle have the potential as a safeguards surveillance and accountancy system. The ICT allows the verification of: fabricator's uranium and plutonium content specifications, shipper/receiver differences between fabricator output and reactor input, reactor plant inventory changes, reprocessing batch specifications and shipper/receiver differences between reactor output and reprocessing plant input. The investigation indicates that there exist predictable functional relationships (i.e. correlations) between isotopic concentrations over a range of burnup. Several cross-correlations serve to establish the initial fuel assembly-averaged compositions. The selection of the more effective correlations will depend not only on the level of reliability of ICT for verification, but also on the capability, accuracy and difficulty of developing measurement methods. The propagation of measurement errors through the correlations have been examined to identify the sensitivity of the isotope correlations to measurement errors, and to establish criteria for measurement accuracy in the development and selection of measurement methods. 6 figures, 3 tables

  10. Pharmacokinetics of lidocaine and bupivacaine and stable isotope labelled analogues : a study in healthy volunteers

    NARCIS (Netherlands)

    Burm, A.G.D.; de Boer, A G; van Kleef, J.W.; Vermeulen, N P; de Leede, L G; Spierdijk, J; Breimer, D D

    1988-01-01

    The pharmacokinetics of lidocaine and bupivacaine and tri-deuteromethyl-labelled lidocaine and bupivacaine were investigated in healthy volunteers. The deuterium-labelled and the unlabelled form of the drug to be investigated were simultaneously infused in 10 min. Plasma concentrations were

  11. Qualitative Metabolome Analysis of Human Cerebrospinal Fluid by 13C-/12C-Isotope Dansylation Labeling Combined with Liquid Chromatography Fourier Transform Ion Cyclotron Resonance Mass Spectrometry

    Science.gov (United States)

    Guo, Kevin; Bamforth, Fiona; Li, Liang

    2011-02-01

    Metabolome analysis of human cerebrospinal fluid (CSF) is challenging because of low abundance of metabolites present in a small volume of sample. We describe and apply a sensitive isotope labeling LC-MS technique for qualitative analysis of the CSF metabolome. After a CSF sample is divided into two aliquots, they are labeled by 13C-dansyl and 12C-dansyl chloride, respectively. The differentially labeled aliquots are then mixed and subjected to LC-MS using Fourier-transform ion cyclotron resonance mass spectrometry (FTICR MS). Dansylation offers significant improvement in the performance of chromatography separation and detection sensitivity. Moreover, peaks detected in the mass spectra can be readily analyzed for ion pair recognition and database search based on accurate mass and/or retention time information. It is shown that about 14,000 features can be detected in a 25-min LC-FTICR MS run of a dansyl-labeled CSF sample, from which about 500 metabolites can be profiled. Results from four CSF samples are compared to gauge the detectability of metabolites by this method. About 261 metabolites are commonly detected in replicate runs of four samples. In total, 1132 unique metabolite ion pairs are detected and 347 pairs (31%) matched with at least one metabolite in the Human Metabolome Database. We also report a dansylation library of 220 standard compounds and, using this library, about 85 metabolites can be positively identified. Among them, 21 metabolites have never been reported to be associated with CSF. These results illustrate that the dansylation LC-FTICR MS method can be used to analyze the CSF metabolome in a more comprehensive manner.

  12. Qualitative metabolome analysis of human cerebrospinal fluid by 13C-/12C-isotope dansylation labeling combined with liquid chromatography Fourier transform ion cyclotron resonance mass spectrometry.

    Science.gov (United States)

    Guo, Kevin; Bamforth, Fiona; Li, Liang

    2011-02-01

    Metabolome analysis of human cerebrospinal fluid (CSF) is challenging because of low abundance of metabolites present in a small volume of sample. We describe and apply a sensitive isotope labeling LC-MS technique for qualitative analysis of the CSF metabolome. After a CSF sample is divided into two aliquots, they are labeled by (13)C-dansyl and (12)C-dansyl chloride, respectively. The differentially labeled aliquots are then mixed and subjected to LC-MS using Fourier-transform ion cyclotron resonance mass spectrometry (FTICR MS). Dansylation offers significant improvement in the performance of chromatography separation and detection sensitivity. Moreover, peaks detected in the mass spectra can be readily analyzed for ion pair recognition and database search based on accurate mass and/or retention time information. It is shown that about 14,000 features can be detected in a 25-min LC-FTICR MS run of a dansyl-labeled CSF sample, from which about 500 metabolites can be profiled. Results from four CSF samples are compared to gauge the detectability of metabolites by this method. About 261 metabolites are commonly detected in replicate runs of four samples. In total, 1132 unique metabolite ion pairs are detected and 347 pairs (31%) matched with at least one metabolite in the Human Metabolome Database. We also report a dansylation library of 220 standard compounds and, using this library, about 85 metabolites can be positively identified. Among them, 21 metabolites have never been reported to be associated with CSF. These results illustrate that the dansylation LC-FTICR MS method can be used to analyze the CSF metabolome in a more comprehensive manner. © American Society for Mass Spectrometry, 2011

  13. Alternative methods of synthesizing 99Tcm-labelled ciprofloxacin

    International Nuclear Information System (INIS)

    Kumar, V.; Choong, K.K.L.; Evans, S.; Olma, T.R.

    1999-01-01

    Full text: 99 Tc m -labelled ciprofloxacin (Infecton) is a new class of radiopharmaceutical designed for imaging live bacterial infection. We synthesized Infecton by modifying the procedure described by Keith Britton's group (Lancet 1996; 347: 233-235) and reported our findings at the ANZSNM meeting last year. Since the methodology was cumbersome, we investigated simpler alternative ways of labelling ciprofloxacin with 99 Tc m -pertechnetate for routine imaging. There were several limitations in the previously described method: (1) Need to prepare pure ciprofloxacin which was unstable on storage. (2) Synthetic procedure using formimidine sulphinic acid (FSA) was complicated and required boiling step. (3) The radiochemical purity (RCP) of the product was low (45-50%) requiring purification. (4) Biodistribution studies showed a marked uptake by the liver which could interfere with scan interpretation in this region. The results of our present studies showed that Infecton could be prepared by a simple two-step method: (1) Reduce 99 Tc m -pertechnetate with stannous salt (SnCl 2 or Sn-tartrate). (2) Mix with Ciproxin IV-100. The RCP of the product was up to 98%, which obviates the need for further purification. Infecton synthesized by the above method showed avid localization in abscesses induced with Staphylococcus aureus in rats. The biodistribution studies showed that Infecton was renally excreted with minimal accumulation in the liver or other organs

  14. From position-specific isotope labeling towards soil fluxomics: a novel toolbox to assess the microbial impact on biogeochemical cycles

    Science.gov (United States)

    Apostel, C.; Dippold, M. A.; Kuzyakov, Y.

    2015-12-01

    Understanding the microbial impact on C and nutrient cycles is one of the most important challenges in terrestrial biogeochemistry. Transformation of low molecular weight organic substances (LMWOS) is a key step in all biogeochemical cycles because 1) all high molecular substances pass the LMWOS pool during their degradation and 2) only LMWOS can be taken up by microorganisms intact. Thus, the transformations of LMWOS are dominated by biochemical pathways of the soil microorganisms. Thus, understanding fluxes and transformations in soils requires a detailed knowledge on the microbial metabolic network and its control mechanism. Tracing C fate in soil by isotopes became on of the most applied and promising biogeochemistry tools but studies were nearly exclusively based on uniformly labeled substances. However, such tracers do not allow the differentiation of the intact use of the initial substances from its transformation to metabolites. The novel tool of position-specific labeling enables to trace molecule atoms separately and thus to determine the cleavage of molecules - a prerequisite for metabolic tracing. Position-specific labeling of basic metabolites and quantification of isotope incorporation in CO2 and bulk soil enabled following the basic metabolic pathways of microorganisms. However, the combination of position-specific 13C labeling with compound-specific isotope analysis of microbial biomarkers and metabolites like phospholipid fatty acids (PLFA) or amino sugars revealed new insights into the soil fluxome: First, it enables tracing specific anabolic pathways in diverse microbial communities in soils e.g. carbon starvation pathways versus pathways reflecting microbial growth. Second, it allows identification of specific pathways of individual functional microbial groups in soils in situ. Tracing metabolic pathways and understanding their regulating factors are crucial for soil C fluxomics i.e. the unravaling of the complex network of C transformations

  15. Microspheres labelled with short-lived isotopes: Development and application for tumors treatment (Experimental study)

    International Nuclear Information System (INIS)

    Drozdovsky, B.Y.; Rosiev, R.A.; Goncharova, A.Y.; Skvortsov, V.G.; Petriev, V.M.; Grigoriev, A.N.; Schischkanov, N.G.

    1997-01-01

    Analysis of the conducted studies strongly suggests the possibility of usage of the domestic protein microspheres as a vehicle for radionuclide. The neutron-activating method of RPP production enables to utilize a broad spectrum of short-living isotopes that can be delivered into the target organ and anchored there for a long time. Good treatment results were obtained in case of the experimentally induced rheumatoid arthritis in rats after intraarticular loading of 165 Dy-hMSA. Mathematical calculations show that homogeneous distribution of RPP in human articulation cavity with the square of 100 cm 2 can be achieved when the quantity of administered particles exceeds 3000. On the example of 165 Dy-hMSA energy characteristic distribution we demonstrated that the absorbed dose for damaged cells at 2mm distance from the radioactive source is 7 times less than the one for a sphere of 2mm diameter. Analysis of dosimetric data in case of intratumoral loading of 165 Dy-hMSA also point out the necessity of the absorbed dose calculation methods taking into account the distance from the source and possible heterogeneity of RPP distribution inside the tumor to be employed. The prolonged RPP detention in the target causing no essential morphological and functional changes was achieved by embolization on the level of septal and interlobular arteries and of efferent arterioles in the animal's renal. The uniformity of microsphere distribution in the organ and their accumulation in tumors depends on the number of particles being administered. Investigations carried out suggest the efficacy of radionuclide therapy application for treatment of oncological and heavy somatic diseases. They also indicate the necessity of further investigations aimed to optimize the usage of microspheres as a radionuclide carrier usage and to work out the criteria of dosimetric planning

  16. Microspheres labelled with short-lived isotopes: Development and application for tumors treatment (Experimental study)

    Energy Technology Data Exchange (ETDEWEB)

    Drozdovsky, B.Y.; Rosiev, R.A.; Goncharova, A.Y.; Skvortsov, V.G.; Petriev, V.M.; Grigoriev, A.N.; Schischkanov, N.G. [Medical Radiological Research Centre RAMS, Kaluga Region, (Russian Federation)

    1997-10-01

    Analysis of the conducted studies strongly suggests the possibility of usage of the domestic protein microspheres as a vehicle for radionuclide. The neutron-activating method of RPP production enables to utilize a broad spectrum of short-living isotopes that can be delivered into the target organ and anchored there for a long time. Good treatment results were obtained in case of the experimentally induced rheumatoid arthritis in rats after intraarticular loading of {sup 165}Dy-hMSA. Mathematical calculations show that homogeneous distribution of RPP in human articulation cavity with the square of 100 cm{sup 2} can be achieved when the quantity of administered particles exceeds 3000. On the example of {sup 165}Dy-hMSA energy characteristic distribution we demonstrated that the absorbed dose for damaged cells at 2mm distance from the radioactive source is 7 times less than the one for a sphere of 2mm diameter. Analysis of dosimetric data in case of intratumoral loading of {sup 165}Dy-hMSA also point out the necessity of the absorbed dose calculation methods taking into account the distance from the source and possible heterogeneity of RPP distribution inside the tumor to be employed. The prolonged RPP detention in the target causing no essential morphological and functional changes was achieved by embolization on the level of septal and interlobular arteries and of efferent arterioles in the animal`s renal. The uniformity of microsphere distribution in the organ and their accumulation in tumors depends on the number of particles being administered. Investigations carried out suggest the efficacy of radionuclide therapy application for treatment of oncological and heavy somatic diseases. They also indicate the necessity of further investigations aimed to optimize the usage of microspheres as a radionuclide carrier usage and to work out the criteria of dosimetric planning 25 refs.

  17. New labeling methods via organometallic species: new synthesis of a chiral methyl group

    International Nuclear Information System (INIS)

    Faucher, Nicolas

    2000-01-01

    Chapter 1: New labeling methods via organometallic species. In the first part of this work, we have developed a new labeling strategy based on the hydrogenolysis of organolithium compounds with tritium gas or deuterium gas. This reaction is catalyzed with palladium on charcoal and leads to the labelled compounds with direct replacement of the proton by its isotopes ("2H or "3H) without further chemical modification of the target molecule. Using this strategy, tritium or deuterium atoms can be introduced in a region but also in a stereoselective fashion with more than 90% ee. The former result was obtained using (-)-sparteine during the lithiation step. Chapter II: New synthesis of a chiral methyl group. In the second part of this work, we have developed a new synthetic method to prepare chiral ditosyl-methylamine using 4,5-disubstituted oxazolidines. Dia-stereoselective substitution of the methoxy group of a 2-alkoxy-oxazolidine by a deuteride in the presence of a Lewis acid leads to the 2-deutero-oxazolidine in a highly stereoselective fashion (de = 100%). Still using a lewis acid, a tritiated hydride open the former 2-deutero-oxazolidine to afford chiral methyl group borne by the nitrogen. Further de-protection and re-protection steps lead to the ditosyl-methylamine with an ee of 65% (RIS= 83/17). Nowadays, this is the best known synthetic method, not only in terms of enantioselectivity but also in terms of chemical yield and number of radioactive steps. As NTs_2 is a fairly good leaving group, the ditosyl-methylamine offers the possibility of introducing chiral methyl group in many substrates using a S_N2 reaction with various nucleophiles. This last point leads to many potential applications in the field of biochemistry or for mechanical studies. (author) [fr

  18. Labelled antibody assays for measuring free triiodothyronine: evaluation and comparison with a labelled analog method

    International Nuclear Information System (INIS)

    Sapin, R.; Gasser, F.; Schlienger, J.L.; Chambron, J.

    1993-01-01

    We evaluated analytically and clinically two new one-step labelled antibody assays for measuring free triiodothyronine (FT3): the first, radiolabelled with 125 I, Amerlex-MAB (MAB) from Kodak diagnostic, and the second, labelled with peroxidase, Enzymum-test FT3 (BM) from Boehringer Mannheim adapted for the Boehringer ES 600 analyzer. The clinical results were compared with those obtained with a radiolabelled analog tracer kit, Amerlex-M (M) from Kodak diagnostic. The latter kit is known to give low FT3 results in sera with low albumin concentrations. Analytical performances of the automated method (BM) were better than those obtained with the manual method (MAB): intra-assay reproducibility (CV<3% vs CV about 5%), inter-assay reproducibility (CV<4% vs CV between 4 and 8%) and mean drift (+1.25% vs -4.3%). The detection limit was low for both kits (<1 pmol/l). In the euthyroid reference group (n = 98) we observed a significant difference between outpatient and hospitalized patient FT3 concentrations as measured with the M kit only. Clinical sensitivity for hyperthyroidism (n = 38) was better for the MAB (92%) than for the BM kit (76%). Specificity in euthyroid L-thyroxine (T4) treated patients (n = 26) was good for both kits (MAB: 92%; BM: 88%) . Hypoalbuminemia (n = 8) decreased FT3 results as follows: M, very significantly; BM, significantly; MAB, only slightly. In patients treated with amiodarone (n = 5), a drug known to lower the metabolic conversion of T4 to T3, the increase of the MAB FT3 results contrasted with the decrease of the BM and M results. In conclusion, results of the two new kits were not strongly influenced by hypoalbuminemia. The MAB results showing lack of decrease in patients with non-thyroidal illness without hypoalbuminemia and in amiodarone-treated patients were unexpected

  19. Efficient method of enzymatic synthesis of nucleosides labelled with 14C and 3H

    International Nuclear Information System (INIS)

    Nejedly, Z.; Filip, J.

    1988-01-01

    The method is presented of enzymatic synthesis of nucleosides labelled with 14 C or 3 H either uniformly or specifically in the base or the deoxyribosyl or ribosyl moiety. The method is based on the ribosylation or deoxyribosylation of the nucleic acid bases (non-labelled or labelled with 14 C or 3 H) by the catalytic effect of enzymes occurring in the supernatant fractions of non-purified homogenates of Escherichia coli B. bacteria. The non-labelled and labelled nucleosides are used as donors of ribosyl or deoxyribosyl groups. The HPLC method is used for separating labelled nucleosides. The radiochemical purity of the labelled nucleosides is higher than 98%, molar activity ranges from 9.2 to 18.5 GBq.mmol -1 ( 14 C-labelled compounds) and from 0.6 to 1.9 TBq.mmol -1 (3H-labelled compounds). (author). 4 figs., 8 refs

  20. Applying quantitative metabolomics based on chemical isotope labeling LC-MS for detecting potential milk adulterant in human milk.

    Science.gov (United States)

    Mung, Dorothea; Li, Liang

    2018-02-25

    There is an increasing demand for donor human milk to feed infants for various reasons including that a mother may be unable to provide sufficient amounts of milk for their child or the milk is considered unsafe for the baby. Selling and buying human milk via the Internet has gained popularity. However, there is a risk of human milk sold containing other adulterants such as animal or plant milk. Analytical tools for rapid detection of adulterants in human milk are needed. We report a quantitative metabolomics method for detecting potential milk adulterants (soy, almond, cow, goat and infant formula milk) in human milk. It is based on the use of a high-performance chemical isotope labeling (CIL) LC-MS platform to profile the metabolome of an unknown milk sample, followed by multivariate or univariate comparison of the resultant metabolomic profile with that of human milk to determine the differences. Using dansylation LC-MS to profile the amine/phenol submetabolome, we could detect an average of 4129 ± 297 (n = 9) soy metabolites, 3080 ± 470 (n = 9) almond metabolites, 4256 ± 136 (n = 18) cow metabolites, 4318 ± 198 (n = 9) goat metabolites, 4444 ± 563 (n = 9) infant formula metabolites, and 4020 ± 375 (n = 30) human metabolites. This high level of coverage allowed us to readily differentiate the six different types of samples. From the analysis of binary mixtures of human milk containing 5, 10, 25, 50 and 75% other type of milk, we demonstrated that this method could be used to detect the presence of as low as 5% adulterant in human milk. We envisage that this method could be applied to detect contaminant or adulterant in other types of food or drinks. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Architecture and dynamics of isotopically labelled macromolecules by nuclear magnetic resonance spectroscopy

    International Nuclear Information System (INIS)

    Matwiyoff, N.A.

    1979-01-01

    The use of 13 C is considered using NMR spectra of cell suspensions. Biochemical reaction kinetics are still unclear in the study of environmental and structural perturbations of amino acids and peptides; thus needs still exist for this labelling technique

  2. Field Sample Preparation Method Development for Isotope Ratio Mass Spectrometry

    International Nuclear Information System (INIS)

    Leibman, C.; Weisbrod, K.; Yoshida, T.

    2015-01-01

    Non-proliferation and International Security (NA-241) established a working group of researchers from Los Alamos National Laboratory (LANL), Pacific Northwest National Laboratory (PNNL) and Savannah River National Laboratory (SRNL) to evaluate the utilization of in-field mass spectrometry for safeguards applications. The survey of commercial off-the-shelf (COTS) mass spectrometers (MS) revealed no instrumentation existed capable of meeting all the potential safeguards requirements for performance, portability, and ease of use. Additionally, fieldable instruments are unlikely to meet the International Target Values (ITVs) for accuracy and precision for isotope ratio measurements achieved with laboratory methods. The major gaps identified for in-field actinide isotope ratio analysis were in the areas of: 1. sample preparation and/or sample introduction, 2. size reduction of mass analyzers and ionization sources, 3. system automation, and 4. decreased system cost. Development work in 2 through 4, numerated above continues, in the private and public sector. LANL is focusing on developing sample preparation/sample introduction methods for use with the different sample types anticipated for safeguard applications. Addressing sample handling and sample preparation methods for MS analysis will enable use of new MS instrumentation as it becomes commercially available. As one example, we have developed a rapid, sample preparation method for dissolution of uranium and plutonium oxides using ammonium bifluoride (ABF). ABF is a significantly safer and faster alternative to digestion with boiling combinations of highly concentrated mineral acids. Actinides digested with ABF yield fluorides, which can then be analyzed directly or chemically converted and separated using established column chromatography techniques as needed prior to isotope analysis. The reagent volumes and the sample processing steps associated with ABF sample digestion lend themselves to automation and field

  3. Combining position-specific 13C labeling with compound-specific isotope analysis: first steps towards soil fluxomics

    Science.gov (United States)

    Dippold, Michaela; Kuzyakov, Yakov

    2015-04-01

    Understanding the soil organic matter (SOM) dynamics is one of the most important challenges in soil science. Transformation of low molecular weight organic substances (LMWOS) is a key step in biogeochemical cycles because 1) all high molecular substances pass this stage during their decomposition and 2) only LMWOS will be taken up by microorganisms. Previous studies on LMWOS were focused on determining net fluxes through the LMWOS pool, but they rarely identified transformations. As LMWOS are the preferred C and energy source for microorganisms, the transformations of LMWOS are dominated by biochemical pathways of the soil microorganisms. Thus, understanding fluxes and transformations in soils requires a detailed knowledge on the biochemical pathways and its controlling factors. Tracing C fate in soil by isotopes became on of the most applied and promising biogeochemistry tools. Up to now, studies on LMWOS were nearly exclusively based on uniformly labeled organic substances i.e. all C atoms in the molecules were labeled with 13C or 14C. However, this classical approach did not allow the differentiation between use of intact initial substances in any process, or whether they were transformed to metabolites. The novel tool of position-specific labeling enables to trace molecule atoms separately and thus to determine the cleavage of molecules - a prerequisite for metabolic tracing. Position-specific labeling of LMWOS and quantification of 13CO2 and 13C in bulk soil enabled following the basic metabolic pathways of soil microorganisms. However, only the combination of position-specific 13C labeling with compound-specific isotope analysis of microbial biomarkers and metabolites allowed 1) tracing specific anabolic pathways in diverse microbial communities in soils and 2) identification of specific pathways of individual functional microbial groups. So, these are the prerequisites for soil fluxomics. Our studies combining position-specific labeled glucose with amino

  4. Measurement of Endogenous versus Exogenous Formaldehyde-Induced DNA-Protein Crosslinks in Animal Tissues by Stable Isotope Labeling and Ultrasensitive Mass Spectrometry.

    Science.gov (United States)

    Lai, Yongquan; Yu, Rui; Hartwell, Hadley J; Moeller, Benjamin C; Bodnar, Wanda M; Swenberg, James A

    2016-05-01

    DNA-protein crosslinks (DPC) arise from a wide range of endogenous and exogenous chemicals, such as chemotherapeutic drugs and formaldehyde. Importantly, recent identification of aldehydes as endogenous genotoxins in Fanconi anemia has provided new insight into disease causation. Because of their bulky nature, DPCs pose severe threats to genome stability, but previous methods to measure formaldehyde-induced DPCs were incapable of discriminating between endogenous and exogenous sources of chemical. In this study, we developed methods that provide accurate and distinct measurements of both exogenous and endogenous DPCs in a structurally specific manner. We exposed experimental animals to stable isotope-labeled formaldehyde ([(13)CD2]-formaldehyde) by inhalation and performed ultrasensitive mass spectrometry to measure endogenous (unlabeled) and exogenous ((13)CD2-labeled) DPCs. We found that exogenous DPCs readily accumulated in nasal respiratory tissues but were absent in tissues distant to the site of contact. This observation, together with the finding that endogenous formaldehyde-induced DPCs were present in all tissues examined, suggests that endogenous DPCs may be responsible for increased risks of bone marrow toxicity and leukemia. Furthermore, the slow rate of DPC repair provided evidence for the persistence of DPCs. In conclusion, our method for measuring endogenous and exogenous DPCs presents a new perspective for the potential health risks inflicted by endogenous formaldehyde and may inform improved disease prevention and treatment strategies. Cancer Res; 76(9); 2652-61. ©2016 AACR. ©2016 American Association for Cancer Research.

  5. Lifetimes of organic photovoltaics: Using TOF-SIMS and 18O2 isotopic labelling to characterise chemical degradation mechanisms

    DEFF Research Database (Denmark)

    Norrman, K.; Krebs, Frederik C

    2006-01-01

    The lifetimes of organic photovoltaic cells based on conjugated polymer materials were studied. The device geometry was glass:ITO:PEDOT:PSS:C-12-PSV:C-60:aluminium. To characterise and elucidate the parts of the degradation mechanisms induced by molecular oxygen, 1802 isotopic labelling was emplo......The lifetimes of organic photovoltaic cells based on conjugated polymer materials were studied. The device geometry was glass:ITO:PEDOT:PSS:C-12-PSV:C-60:aluminium. To characterise and elucidate the parts of the degradation mechanisms induced by molecular oxygen, 1802 isotopic labelling...

  6. Probing the metabolic network in bloodstream-form Trypanosoma brucei using untargeted metabolomics with stable isotope labelled glucose.

    Directory of Open Access Journals (Sweden)

    Darren J Creek

    2015-03-01

    Full Text Available Metabolomics coupled with heavy-atom isotope-labelled glucose has been used to probe the metabolic pathways active in cultured bloodstream form trypomastigotes of Trypanosoma brucei, a parasite responsible for human African trypanosomiasis. Glucose enters many branches of metabolism beyond glycolysis, which has been widely held to be the sole route of glucose metabolism. Whilst pyruvate is the major end-product of glucose catabolism, its transamination product, alanine, is also produced in significant quantities. The oxidative branch of the pentose phosphate pathway is operative, although the non-oxidative branch is not. Ribose 5-phosphate generated through this pathway distributes widely into nucleotide synthesis and other branches of metabolism. Acetate, derived from glucose, is found associated with a range of acetylated amino acids and, to a lesser extent, fatty acids; while labelled glycerol is found in many glycerophospholipids. Glucose also enters inositol and several sugar nucleotides that serve as precursors to macromolecule biosynthesis. Although a Krebs cycle is not operative, malate, fumarate and succinate, primarily labelled in three carbons, were present, indicating an origin from phosphoenolpyruvate via oxaloacetate. Interestingly, the enzyme responsible for conversion of phosphoenolpyruvate to oxaloacetate, phosphoenolpyruvate carboxykinase, was shown to be essential to the bloodstream form trypanosomes, as demonstrated by the lethal phenotype induced by RNAi-mediated downregulation of its expression. In addition, glucose derivatives enter pyrimidine biosynthesis via oxaloacetate as a precursor to aspartate and orotate.

  7. A Convenient Method for Estimation of the Isotopic Abundance in Uranium Bearing Samples

    International Nuclear Information System (INIS)

    AI -Saleh, F.S.; AI-Mukren, Alj.H.; Farouk, M.A.

    2008-01-01

    A convenient and simple method for estimation of the isotopic abundance in some uranium bearing samples using gamma-ray spectrometry is developed using a hyper pure germanium spectrometer and a standard uranium sample with known isotopic abundance

  8. Calcium Isotope Analysis with "Peak Cut" Method on Column Chemistry

    Science.gov (United States)

    Zhu, H.; Zhang, Z.; Liu, F.; Li, X.

    2017-12-01

    To eliminate isobaric interferences from elemental and molecular isobars (e.g., 40K+, 48Ti+, 88Sr2+, 24Mg16O+, 27Al16O+) on Ca isotopes during mass determination, samples should be purified through ion-exchange column chemistry before analysis. However, large Ca isotopic fractionation has been observed during column chemistry (Russell and Papanastassiou, 1978; Zhu et al., 2016). Therefore, full recovery during column chemistry is greatly needed, otherwise uncertainties would be caused by poor recovery (Zhu et al., 2016). Generally, matrix effects could be enhanced by full recovery, as other elements might overlap with Ca cut during column chemistry. Matrix effects and full recovery are difficult to balance and both need to be considered for high-precision analysis of stable Ca isotopes. Here, we investigate the influence of poor recovery on δ44/40Ca using TIMS with the double spike technique. The δ44/40Ca values of IAPSO seawater, ML3B-G and BHVO-2 in different Ca subcats (e.g., 0-20, 20-40, 40-60, 60-80, 80-100%) with 20% Ca recovery on column chemistry display limited variation after correction by the 42Ca-43Ca double spike technique with the exponential law. Notably, δ44/40Ca of each Ca subcut is quite consistent with δ44/40Ca of Ca cut with full recovery within error. Our results indicate that the 42Ca-43Ca double spike technique can simultaneously correct both of the Ca isotopic fractionation that occurred during column chemistry and thermal ionization mass spectrometry (TIMS) determination properly, because both of the isotopic fractionation occurred during analysis follow the exponential law well. Therefore, we propose the "peak cut" method on Ca column chemistry for samples with complex matrix effects. Briefly, for samples with low Ca contents, we can add the double spike before column chemistry, and only collect the middle of the Ca eluate and abandon the both sides of Ca eluate that might overlap with other elements (e.g., K, Sr). This method would

  9. Quantitative Isotope-Dilution High-Resolution-Mass-Spectrometry Analysis of Multiple Intracellular Metabolites in Clostridium autoethanogenum with Uniformly 13C-Labeled Standards Derived from Spirulina.

    Science.gov (United States)

    Schatschneider, Sarah; Abdelrazig, Salah; Safo, Laudina; Henstra, Anne M; Millat, Thomas; Kim, Dong-Hyun; Winzer, Klaus; Minton, Nigel P; Barrett, David A

    2018-04-03

    We have investigated the applicability of commercially available lyophilized spirulina ( Arthrospira platensis), a microorganism uniformly labeled with 13 C, as a readily accessible source of multiple 13 C-labeled metabolites suitable as internal standards for the quantitative determination of intracellular bacterial metabolites. Metabolites of interest were analyzed by hydrophilic-interaction liquid chromatography coupled with high-resolution mass spectrometry. Multiple internal standards obtained from uniformly (U)- 13 C-labeled extracts from spirulina were used to enable isotope-dilution mass spectrometry (IDMS) in the identification and quantification of intracellular metabolites. Extraction of the intracellular metabolites of Clostridium autoethanogenum using 2:1:1 chloroform/methanol/water was found to be the optimal method in comparison with freeze-thaw, homogenization, and sonication methods. The limits of quantification were ≤1 μM with excellent linearity for all of the calibration curves ( R 2 ≥ 0.99) for 74 metabolites. The precision and accuracy were found to be within relative standard deviations (RSDs) of 15% for 49 of the metabolites and within RSDs of 20% for all of the metabolites. The method was applied to study the effects of feeding different levels of carbon monoxide (as a carbon source) on the central metabolism and Wood-Ljungdahl pathway of C. autoethanogenum grown in continuous culture over 35 days. Using LC-IDMS with U- 13 C spirulina allowed the successful quantification of 52 metabolites in the samples, including amino acids, carboxylic acids, sugar phosphates, purines, and pyrimidines. The method provided absolute quantitative data on intracellular metabolites that was suitable for computational modeling to understand and optimize the C. autoethanogenum metabolic pathways active in gas fermentation.

  10. Assay for vitamin B12 absorption and method of making labeled vitamin B12

    Science.gov (United States)

    Anderson, Peter J [Davis, CA; Dueker, Stephen [Davis, CA; Miller, Joshua [Davis, CA; Green, Ralph [Elmacero, CA; Roth, John [Davis, CA; Carkeet, Colleen [Silver Spring, MD; Buchholz,; Bruce, A [Orinda, CA

    2012-06-19

    The invention provides methods for labeling vitamin B12 with .sup.14C, .sup.13C, tritium, and deuterium. When radioisotopes are used, the invention provides for methods of labeling B12 with high specific activity. The invention also provides labeled vitamin B12 compositions made in accordance with the invention.

  11. Automated NMR structure determination of stereo-array isotope labeled ubiquitin from minimal sets of spectra using the SAIL-FLYA system

    Energy Technology Data Exchange (ETDEWEB)

    Ikeya, Teppei [Goethe University Frankfurt am Main, Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance (Germany); Takeda, Mitsuhiro; Yoshida, Hitoshi; Terauchi, Tsutomu; Jee, Jun-Goo; Kainosho, Masatsune [Tokyo Metropolitan University, Graduate School of Science (Japan)], E-mail: kainosho@nmr.chem.metro-u.ac.jp; Guentert, Peter [Goethe University Frankfurt am Main, Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance (Germany)], E-mail: guentert@em.uni-frankfurt.de

    2009-08-15

    Stereo-array isotope labeling (SAIL) has been combined with the fully automated NMR structure determination algorithm FLYA to determine the three-dimensional structure of the protein ubiquitin from different sets of input NMR spectra. SAIL provides a complete stereo- and regio-specific pattern of stable isotopes that results in sharper resonance lines and reduced signal overlap, without information loss. Here we show that as a result of the superior quality of the SAIL NMR spectra, reliable, fully automated analyses of the NMR spectra and structure calculations are possible using fewer input spectra than with conventional uniformly {sup 13}C/{sup 15}N-labeled proteins. FLYA calculations with SAIL ubiquitin, using a single three-dimensional 'through-bond' spectrum (and 2D HSQC spectra) in addition to the {sup 13}C-edited and {sup 15}N-edited NOESY spectra for conformational restraints, yielded structures with an accuracy of 0.83-1.15 A for the backbone RMSD to the conventionally determined solution structure of SAIL ubiquitin. NMR structures can thus be determined almost exclusively from the NOESY spectra that yield the conformational restraints, without the need to record many spectra only for determining intermediate, auxiliary data of the chemical shift assignments. The FLYA calculations for this report resulted in 252 ubiquitin structure bundles, obtained with different input data but identical structure calculation and refinement methods. These structures cover the entire range from highly accurate structures to seriously, but not trivially, wrong structures, and thus constitute a valuable database for the substantiation of structure validation methods.

  12. Automated NMR structure determination of stereo-array isotope labeled ubiquitin from minimal sets of spectra using the SAIL-FLYA system

    International Nuclear Information System (INIS)

    Ikeya, Teppei; Takeda, Mitsuhiro; Yoshida, Hitoshi; Terauchi, Tsutomu; Jee, Jun-Goo; Kainosho, Masatsune; Guentert, Peter

    2009-01-01

    Stereo-array isotope labeling (SAIL) has been combined with the fully automated NMR structure determination algorithm FLYA to determine the three-dimensional structure of the protein ubiquitin from different sets of input NMR spectra. SAIL provides a complete stereo- and regio-specific pattern of stable isotopes that results in sharper resonance lines and reduced signal overlap, without information loss. Here we show that as a result of the superior quality of the SAIL NMR spectra, reliable, fully automated analyses of the NMR spectra and structure calculations are possible using fewer input spectra than with conventional uniformly 13 C/ 15 N-labeled proteins. FLYA calculations with SAIL ubiquitin, using a single three-dimensional 'through-bond' spectrum (and 2D HSQC spectra) in addition to the 13 C-edited and 15 N-edited NOESY spectra for conformational restraints, yielded structures with an accuracy of 0.83-1.15 A for the backbone RMSD to the conventionally determined solution structure of SAIL ubiquitin. NMR structures can thus be determined almost exclusively from the NOESY spectra that yield the conformational restraints, without the need to record many spectra only for determining intermediate, auxiliary data of the chemical shift assignments. The FLYA calculations for this report resulted in 252 ubiquitin structure bundles, obtained with different input data but identical structure calculation and refinement methods. These structures cover the entire range from highly accurate structures to seriously, but not trivially, wrong structures, and thus constitute a valuable database for the substantiation of structure validation methods

  13. Automated NMR structure determination of stereo-array isotope labeled ubiquitin from minimal sets of spectra using the SAIL-FLYA system.

    Science.gov (United States)

    Ikeya, Teppei; Takeda, Mitsuhiro; Yoshida, Hitoshi; Terauchi, Tsutomu; Jee, Jun-Goo; Kainosho, Masatsune; Güntert, Peter

    2009-08-01

    Stereo-array isotope labeling (SAIL) has been combined with the fully automated NMR structure determination algorithm FLYA to determine the three-dimensional structure of the protein ubiquitin from different sets of input NMR spectra. SAIL provides a complete stereo- and regio-specific pattern of stable isotopes that results in sharper resonance lines and reduced signal overlap, without information loss. Here we show that as a result of the superior quality of the SAIL NMR spectra, reliable, fully automated analyses of the NMR spectra and structure calculations are possible using fewer input spectra than with conventional uniformly 13C/15N-labeled proteins. FLYA calculations with SAIL ubiquitin, using a single three-dimensional "through-bond" spectrum (and 2D HSQC spectra) in addition to the 13C-edited and 15N-edited NOESY spectra for conformational restraints, yielded structures with an accuracy of 0.83-1.15 A for the backbone RMSD to the conventionally determined solution structure of SAIL ubiquitin. NMR structures can thus be determined almost exclusively from the NOESY spectra that yield the conformational restraints, without the need to record many spectra only for determining intermediate, auxiliary data of the chemical shift assignments. The FLYA calculations for this report resulted in 252 ubiquitin structure bundles, obtained with different input data but identical structure calculation and refinement methods. These structures cover the entire range from highly accurate structures to seriously, but not trivially, wrong structures, and thus constitute a valuable database for the substantiation of structure validation methods.

  14. Labeling and stability of radiolabeled antibody fragments by a direct 99mTc-labeling method

    International Nuclear Information System (INIS)

    Pak, K.Y.; Nedelman, M.A.; Tam, S.H.; Wilson, E.; Daddona, P.E.

    1992-01-01

    The in vitro labeling and stability of 99m Tc-labeled antibody Fab' fragments prepared by a direct labeling technique were evaluated. Eight antibody fragments derived from murine IgG1 (N = 5), IgG2a (N = 2) and IgG3 (N = 1) isotypes were labeled with a preformed 99m Tc-D-glucarate complex. No loss of radioactivity incorporation was observed for all the 99m Tc-labeled antibody fragments after 24 h incubation at 37 o C. 99m Tc-labeled antibody fragments (IgG1, N = 2; IgG2a, n = 2; IgG3, N = 1) were stable upon challenge with DTPA, EDTA or acidic pH. Using the affinity chromatography technique, two of the 99m Tc-labeled antibody fragments displayed no loss of immunoreactivity after prolonged incubation in phosphate buffer up to 24 h at 37 o C. Bonding between 99m Tc and antibody fragments was elucidated by challenging with a diamide ditholate (N 2 S 2 ) compound. The Fab' with IgG2a isotype displayed tighter binding to 99m Tc in comparison to Fab' from IgG1 and IgG3 isotype in N 2 S 2 challenge and incubation with human plasma. The in vivo biodistribution of five 99m Tc-labeled fragments were evaluated in normal mice. (Author)

  15. Comparative analysis of monoclonal antibody N-glycosylation using stable isotope labelling and UPLC-fluorescence-MS.

    Science.gov (United States)

    Millán Martín, Silvia; Delporte, Cédric; Farrell, Amy; Navas Iglesias, Natalia; McLoughlin, Niaobh; Bones, Jonathan

    2015-03-07

    A twoplex method using (12)C6 and (13)C6 stable isotope analogues (Δmass = 6 Da) of 2-aminobenzoic acid (2-AA) is described for quantitative analysis of N-glycans present on monoclonal antibodies and other glycoproteins using ultra performance liquid chromatography with sequential fluorescence and accurate mass tandem quadrupole time of flight (QToF) mass spectrometric detection.

  16. IL-2 labeled with 99mTechnetium by an indirect method

    International Nuclear Information System (INIS)

    Bocco, R.; Obenaus, Esteban; Rabiller, Graciela; Castiglia, Silvia G. de

    2003-01-01

    IL-2 and the other cytokines labeled with 99m Tc are an interesting option to early diagnosis of autoimmune diseases and monitoring with nuclear medicine images. The aim of this study was to obtain by indirect method IL-2 labeled with 99m Technetium using Benzoyl MAG3 chelating agent, for in vivo diagnosis of lymphocytic infiltration. IL-2 is a small, relatively fragile protein, and it is essential to retain its receptor binding capacity after labeling. Two different methods of labeling have been proven: Pre-conjugation labeling method: we used NHS- Hynic as a chelator agent and labeled this conjugated protein with 99m Tc using tricine as coligand. Albumin was used as a model for the conjugation and labeling steps. The albumin was labeled by this method with a good labeling efficiency but the IL-2 protein could not be labeled with this approach. Post- conjugation labeling method: we used the bifunctional chelating agent, benzoyl MAG3, this ligand is first labeled with 99m Tc and then is conjugated to the protein. The N 3 S Logan complex was incubated for 30 minutes and then measured by RP- HPLC. An active ester of the labeled Logan was formed and was incubated with Il-2 at room temperature and basic pH to promote the conjugation between the active ester and the protein. The labeling efficiency was determined by RP-HPLC using a C 18 column. The albumin protein was also labeled by this method and the radiochemical purity was measured by RP- HPLC using a GPC column and the labeling efficiency was 80%. The next objectives are to explore different strategies for purifying the 99m Tc-IL-2 and to evaluate the capacity of IL-2 to bind to its receptor after labeling. (author)

  17. Quantitative twoplex glycan analysis using 12C6 and 13C6 stable isotope 2-aminobenzoic acid labelling and capillary electrophoresis mass spectrometry.

    Science.gov (United States)

    Váradi, Csaba; Mittermayr, Stefan; Millán-Martín, Silvia; Bones, Jonathan

    2016-12-01

    Capillary electrophoresis (CE) offers excellent efficiency and orthogonality to liquid chromatographic (LC) separations for oligosaccharide structural analysis. Combination of CE with high resolution mass spectrometry (MS) for glycan analysis remains a challenging task due to the MS incompatibility of background electrolyte buffers and additives commonly used in offline CE separations. Here, a novel method is presented for the analysis of 2-aminobenzoic acid (2-AA) labelled glycans by capillary electrophoresis coupled to mass spectrometry (CE-MS). To ensure maximum resolution and excellent precision without the requirement for excessive analysis times, CE separation conditions including the concentration and pH of the background electrolyte, the effect of applied pressure on the capillary inlet and the capillary length were evaluated. Using readily available 12/13 C 6 stable isotopologues of 2-AA, the developed method can be applied for quantitative glycan profiling in a twoplex manner based on the generation of extracted ion electropherograms (EIE) for 12 C 6 'light' and 13 C 6 'heavy' 2-AA labelled glycan isotope clusters. The twoplex quantitative CE-MS glycan analysis platform is ideally suited for comparability assessment of biopharmaceuticals, such as monoclonal antibodies, for differential glycomic analysis of clinical material for potential biomarker discovery or for quantitative microheterogeneity analysis of different glycosylation sites within a glycoprotein. Additionally, due to the low injection volume requirements of CE, subsequent LC-MS analysis of the same sample can be performed facilitating the use of orthogonal separation techniques for structural elucidation or verification of quantitative performance.

  18. Isotopically labeled sulfur compounds and synthetic selenium and tellurium analogues to study sulfur metabolism in marine bacteria

    Directory of Open Access Journals (Sweden)

    Nelson L. Brock

    2013-05-01

    Full Text Available Members of the marine Roseobacter clade can degrade dimethylsulfoniopropionate (DMSP via competing pathways releasing either methanethiol (MeSH or dimethyl sulfide (DMS. Deuterium-labeled [2H6]DMSP and the synthetic DMSP analogue dimethyltelluriopropionate (DMTeP were used in feeding experiments with the Roseobacter clade members Phaeobacter gallaeciensis DSM 17395 and Ruegeria pomeroyi DSS-3, and their volatile metabolites were analyzed by closed-loop stripping and solid-phase microextraction coupled to GC–MS. Feeding experiments with [2H6]DMSP resulted in the incorporation of a deuterium label into MeSH and DMS. Knockout of relevant genes from the known DMSP demethylation pathway to MeSH showed in both species a residual production of [2H3]MeSH, suggesting that a second demethylation pathway is active. The role of DMSP degradation pathways for MeSH and DMS formation was further investigated by using the synthetic analogue DMTeP as a probe in feeding experiments with the wild-type strain and knockout mutants. Feeding of DMTeP to the R. pomeroyi knockout mutant resulted in a diminished, but not abolished production of demethylation pathway products. These results further corroborated the proposed second demethylation activity in R. pomeroyi. Isotopically labeled [2H3]methionine and 34SO42−, synthesized from elemental 34S8, were tested to identify alternative sulfur sources besides DMSP for the MeSH production in P. gallaeciensis. Methionine proved to be a viable sulfur source for the MeSH volatiles, whereas incorporation of labeling from sulfate was not observed. Moreover, the utilization of selenite and selenate salts by marine alphaproteobacteria for the production of methylated selenium volatiles was explored and resulted in the production of numerous methaneselenol-derived volatiles via reduction and methylation. The pathway of selenate/selenite reduction, however, proved to be strictly separated from sulfate reduction.

  19. Calculation of isotope selective excitation of uranium isotopes using spectral simulation method

    International Nuclear Information System (INIS)

    Al-Hassanieh, O.

    2009-06-01

    Isotope ratio enhancement factor and isotope selectivity of 235 U in five excitation schemes (I: 0→10069 cm - 1 →IP, II: 0 →10081 cm - 1 →IP, III: 0 →25349 cm - 1→ IP, IV: 0→28650 cm - 1 →IP, V: 0→16900 cm - 1 →34659 cm - 1 →IP), were computed by a spectral simulation approach. The effect of laser bandwidth and Doppler width on the isotope ratio enhancement factor and isotope selectivity of 235 U has been studied. The photoionization scheme V gives the highest isotope ratio enhancement factor. The main factors which effect the separation possibility are the isotope shift and the relative intensity of the transitions between hyperfine levels. The isotope ratio enhancement factor decreases exponentially by increasing the Doppler width and the laser bandwidth, where the effect of Doppler width is much greater than the effect of the laser bandwidth. (author)

  20. Analysis of a benthic community food web using isotopically labelled potential food

    International Nuclear Information System (INIS)

    Beviss-Challinor, M.H.; Field, J.G.

    1982-01-01

    A series of experiments was designed to reveal the trophic structure of a benthic community using kelp holdfasts as microcosms within the kelp-bed community. The experimental food comprised zooplankton represented by 3 H 2 O-labelled Artemia sp. eggs and nauplii (200 to 300 μm), detritus derived from 14 C-labelled kelp (60 to 90 μm), and phytoplankton represented by 14 C-labelled Dunaliella primolecta (5 to 10 μm) cultures. Separate experiments of short duration (1 to 2 h) were used to indicate the primary consumers on each type of food, while longer experiments (4, 8 and 16 h) suggested the secondary consumers (coprophagous and carnivorous species). Several species were found to be omnivorous. (orig.)

  1. Synthesis of stereoarray isotope labeled (SAIL) lysine via the "head-to-tail" conversion of SAIL glutamic acid.

    Science.gov (United States)

    Terauchi, Tsutomu; Kamikawai, Tomoe; Vinogradov, Maxim G; Starodubtseva, Eugenia V; Takeda, Mitsuhiro; Kainosho, Masatsune

    2011-01-07

    A stereoarray isotope labeled (SAIL) lysine, (2S,3R,4R,5S,6R)-[3,4,5,6-(2)H(4);1,2,3,4,5,6-(13)C(6);2,6-(15)N(2)]lysine, was synthesized by the "head-to-tail" conversion of SAIL-Glu, (2S,3S,4R)-[3,4-(2)H(2);1,2,3,4,5-(13)C(5);2-(15)N]glutamic acid, with high stereospecificities for all five chiral centers. With the SAIL-Lys in hand, the unambiguous simultaneous stereospecific assignments were able to be established for each of the prochiral protons within the four methylene groups of the Lys side chains in proteins.

  2. Comparative determination of sucrose content in sugar beet by polarimetric and isotope dilution methods

    Energy Technology Data Exchange (ETDEWEB)

    Malec, K; Szuchnik, A [Institute of Nuclear Research, Warsaw (Poland); Rydel, S; Walerianaczyk, E [Instytut Przemyslu Cukrowniczego, Warsaw (Poland)

    1976-01-01

    The comparative determination of sucrose content in sugar beets has been investigated by following methods: polarimetric, direct isotope dilution and double carrier-isotope dilution analysis. Basing upon the obtained results it has been ascertained, that in the case of worse quality beets the polarimetric determinations differ greatly from isotopic data.

  3. Automatic isotope gas analysis of tritium labelled organic materials Pt. 3

    International Nuclear Information System (INIS)

    Gacs, I.; Mlinko, S.; Payer, K.; Otvos, L.; Banfi, D.; Palagyi, T.

    1978-01-01

    An isotope analytical procedure and an automatic instrument developed for the determination of tritium in organic compounds and biological materials by internal gas counting are described. The sample is burnt in a stream of oxygen and the combustion products including water vapour carrying the tritium are led onto a column of molecular sieve-5A heated to 550 deg C. Tritium is retained temporarily on the column, then transferred into a stream of hydrogen by isotope exchange. After addition of butane, the tritiated hydrogen is led into an internal detector and enclosed there for radioactivity measurement. The procedure, providing quantitative recovery, is completed in five minutes. It is free of memory effect and suitable for the determination of tritium in a wide range of organic compounds and samples of biological origin. (author)

  4. Deciphering systemic wound responses of the pumpkin extrafascicular phloem by metabolomics and stable isotope-coded protein labeling.

    Science.gov (United States)

    Gaupels, Frank; Sarioglu, Hakan; Beckmann, Manfred; Hause, Bettina; Spannagl, Manuel; Draper, John; Lindermayr, Christian; Durner, Jörg

    2012-12-01

    In cucurbits, phloem latex exudes from cut sieve tubes of the extrafascicular phloem (EFP), serving in defense against herbivores. We analyzed inducible defense mechanisms in the EFP of pumpkin (Cucurbita maxima) after leaf damage. As an early systemic response, wounding elicited transient accumulation of jasmonates and a decrease in exudation probably due to partial sieve tube occlusion by callose. The energy status of the EFP was enhanced as indicated by increased levels of ATP, phosphate, and intermediates of the citric acid cycle. Gas chromatography coupled to mass spectrometry also revealed that sucrose transport, gluconeogenesis/glycolysis, and amino acid metabolism were up-regulated after wounding. Combining ProteoMiner technology for the enrichment of low-abundance proteins with stable isotope-coded protein labeling, we identified 51 wound-regulated phloem proteins. Two Sucrose-Nonfermenting1-related protein kinases and a 32-kD 14-3-3 protein are candidate central regulators of stress metabolism in the EFP. Other proteins, such as the Silverleaf Whitefly-Induced Protein1, Mitogen Activated Protein Kinase6, and Heat Shock Protein81, have known defensive functions. Isotope-coded protein labeling and western-blot analyses indicated that Cyclophilin18 is a reliable marker for stress responses of the EFP. As a hint toward the induction of redox signaling, we have observed delayed oxidation-triggered polymerization of the major Phloem Protein1 (PP1) and PP2, which correlated with a decline in carbonylation of PP2. In sum, wounding triggered transient sieve tube occlusion, enhanced energy metabolism, and accumulation of defense-related proteins in the pumpkin EFP. The systemic wound response was mediated by jasmonate and redox signaling.

  5. Evaluation of stable isotope labelling strategies for the quantitation of CP4 EPSPS in genetically modified soya

    Energy Technology Data Exchange (ETDEWEB)

    Ocana, Mireia Fernandez [Centre for Chemical and Bioanalytical Sciences, Royal Holloway, University of London, Egham TW20 0EX (United Kingdom)], E-mail: Mireia.FernandezOcana@pfizer.com; Fraser, Paul D. [Centre for Chemical and Bioanalytical Sciences, Royal Holloway, University of London, Egham TW20 0EX (United Kingdom); Patel, Raj K.P.; Halket, John M. [Specialist Bioanalytical Services Ltd., Royal Holloway, University of London, Egham TW20 0EX (United Kingdom); Bramley, Peter M. [Centre for Chemical and Bioanalytical Sciences, Royal Holloway, University of London, Egham TW20 0EX (United Kingdom)

    2009-02-16

    The introduction of genetically modified (GM) crops into the market has raised a general alertness relating to the control and safety of foods. The applicability of protein separation hyphenated to mass spectrometry to identify the bacterial enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS) protein expressed in GM crops has been previously reported [M.F. Ocana, P.D. Fraser, R.K.P. Patel, J.M. Halket, P.M. Bramley, Rapid Commun. Mass Spectrom. 21 (2007) 319.]. Herein, we investigate the suitability of two strategies that employ heavy stable isotopes, i.e. AQUA and iTRAQ, to quantify different levels of CP4 EPSPS in up to four GM preparations. Both quantification strategies showed potential to determine whether the presence of GM material is above the limits established by the European Union. The AQUA quantification procedure involved protein solubilisation/fractionation and subsequent separation using SDS-PAGE. A segment of the gel in which the protein of interest was located was excised, the stable isotope labeled peptide added at a known concentration and proteolytic digestion initiated. Following recovery of the peptides, on-line separation and detection using LC-MS was carried out. A similar approach was used for the iTRAQ workflow with the exception that proteins were digested in solution and generated tryptic peptides were chemically tagged. Both procedures demonstrated the potential for quantitative detection at 0.5% (w/w) GM soya which is a level below the current European Union's threshold for food-labelling. In this context, a comparison between the two procedures is provided within the present study.

  6. Evaluation of stable isotope labelling strategies for the quantitation of CP4 EPSPS in genetically modified soya

    International Nuclear Information System (INIS)

    Ocana, Mireia Fernandez; Fraser, Paul D.; Patel, Raj K.P.; Halket, John M.; Bramley, Peter M.

    2009-01-01

    The introduction of genetically modified (GM) crops into the market has raised a general alertness relating to the control and safety of foods. The applicability of protein separation hyphenated to mass spectrometry to identify the bacterial enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS) protein expressed in GM crops has been previously reported [M.F. Ocana, P.D. Fraser, R.K.P. Patel, J.M. Halket, P.M. Bramley, Rapid Commun. Mass Spectrom. 21 (2007) 319.]. Herein, we investigate the suitability of two strategies that employ heavy stable isotopes, i.e. AQUA and iTRAQ, to quantify different levels of CP4 EPSPS in up to four GM preparations. Both quantification strategies showed potential to determine whether the presence of GM material is above the limits established by the European Union. The AQUA quantification procedure involved protein solubilisation/fractionation and subsequent separation using SDS-PAGE. A segment of the gel in which the protein of interest was located was excised, the stable isotope labeled peptide added at a known concentration and proteolytic digestion initiated. Following recovery of the peptides, on-line separation and detection using LC-MS was carried out. A similar approach was used for the iTRAQ workflow with the exception that proteins were digested in solution and generated tryptic peptides were chemically tagged. Both procedures demonstrated the potential for quantitative detection at 0.5% (w/w) GM soya which is a level below the current European Union's threshold for food-labelling. In this context, a comparison between the two procedures is provided within the present study

  7. Demonstration of de novo synthesis of enzymes by density labelling with stable isotopes

    International Nuclear Information System (INIS)

    Huebner, G.; Hirschberg, K.

    1977-01-01

    The technique of in vivo density labelling of proteins with H 2 18 O and 2 H 2 O has been used to investigate hormonal regulation and developmental expression of enzymes in plant cells. Buoyant density data obtained from isopycnic equilibrium centrifugation demonstrated that the cytokinine-induced nitrate reductase activity and the gibberellic acid-induced phosphatase activity in isolated embryos of Agrostemma githago are activities of enzymes synthesized de novo. The increase in alanine-specific aminopeptidase in germinating A. githago seeds is not due to de novo synthesis but to the release of preformed enzyme. On the basis of this result it is possible to apply the enzyme aminopeptidase as an internal density standard in equilibrium centrifugation. Density labelling experiments on proteins in pea cotyledons have been used to study the change in the activity of acid phosphatase, alanine-specific aminopeptidase, and peroxidase during germination. The activities of these enzymes increase in cotyledons of Pisum sativum. Density labelling by 18 O and 2 H demonstrates de novo synthesis of these three enzymes. The differential time course of enzyme induction shows the advantage of using H 2 18 O as labelling substance in cases when the enzyme was synthesized immediately at the beginning of germination. At this stage of development the amino-acid pool available for synthesis is formed principally by means of hydrolysis of storage proteins. The incorporation of 2 H into the new proteins takes place in a measurable amount at a stage of growth in which the amino acids are also synthesized de novo. The enzyme acid phosphatase of pea cotyledons was chosen to demonstrate the possibility of using the density labelling technique to detect protein turnover. (author)

  8. Dynamics of N2O production pathways analyzed by 15N18O isotope labeling

    DEFF Research Database (Denmark)

    Jensen, Marlene Mark; Ma, Chun; Lavik, Gaute

    Nitrous oxide production associated with biological nitrogen transformations can contribute substantially to the CO2 footprint of both man-made and natural systems, but the pathways and regulation of N2O production are poorly understood. We developed a 15N/18O dual isotope labelling technique...

  9. Stable isotope phenotyping via cluster analysis of NanoSIMS data as a method for characterizing distinct microbial ecophysiologies and sulfur-cycling in the environment

    Directory of Open Access Journals (Sweden)

    Katherine S Dawson

    2016-05-01

    Full Text Available Stable isotope probing (SIP is a valuable tool for gaining insights into ecophysiology and biogeochemical cycling of environmental microbial communities by tracking isotopically labeled compounds into cellular macromolecules as well as into byproducts of respiration. SIP, in conjunction with nanoscale secondary ion mass spectrometry (NanoSIMS, allows for the visualization of isotope incorporation at the single cell level. In this manner, both active cells within a diverse population as well as heterogeneity in metabolism within a homogeneous population can be observed. The ecophysiological implications of these single cell stable isotope measurements are often limited to the taxonomic resolution of paired fluorescence in situ hybridization (FISH microscopy. Here we introduce a taxonomy-independent method using multi-isotope SIP and NanoSIMS for identifying and grouping phenotypically similar microbial cells by their chemical and isotopic fingerprint. This method was applied to SIP experiments in a sulfur-cycling biofilm collected from sulfidic intertidal vents amended with 13C-acetate, 15N-ammonium, and 33S-sulfate. Using a cluster analysis technique based on fuzzy c-means to group cells according to their isotope (13C/12C, 15N/14N, and 33S/32S and elemental ratio (C/CN and S/CN profiles, our analysis partitioned ~2200 cellular regions of interest (ROIs into 5 distinct groups. These isotope phenotype groupings are reflective of the variation in labeled substrate uptake by cells in a multispecies metabolic network dominated by Gamma- and Deltaproteobacteria. Populations independently grouped by isotope phenotype were subsequently compared with paired FISH data, demonstrating a single coherent deltaproteobacterial cluster and multiple gammaproteobacterial groups, highlighting the distinct ecophysiologies of spatially-associated microbes within the sulfur-cycling biofilm from White Point Beach, CA.

  10. Development of method of tritium labeling of pharmacological preparate of drotaverine hydrochloride (NOSPA)

    International Nuclear Information System (INIS)

    Kim, A.A.; Djuraeva, G.T.; Shukurov, B.V.

    2004-01-01

    Full text: The method for tritium labeling of pharmacological preparate of drotaverine hydrochloride (no spa) was developed. Drotaverine hydrochloride was labeled by thermally activated tritium in apparatus for tritium labeling. The optimum regime of labeling was selected. The system of purification of tritium labeled drotaverine hydrochloride by thin layer chromatography (TLC) has been developed. The TLC system of purification of tritium labeled drotaverine hydrochloride was developed. Tritium labeled preparation of drotaverine hydrochloride was purified by TLC on silicagel in system isopropanol: ammonia: water (8:1:1). We found appearance of additional fractions in tritium labeled preparation of drotaverine hydrochloride that testifies to partial transformation of drotaverine hydrochloride during procedure of labeling. Application of TLC for purification of tritium labeled preparation allows to purify completely drotaverine hydrochloride of by-products. The output of purified tritium labeled preparation of drotaverine hydrochloride was about 25 %. The received preparation had specific radioactivity - 3,2 MBq/mg, radiochemical purity of a preparation was 95 %. TLC purification seems inexpensive, fast and suitable for purification of tritium-labeled drotaverine hydrochloride. Thus developed method allows obtain tritium labeled preparation of drotaverine hydrochloride (no - spa), suitable for medical and biologic researches

  11. Isotopic evaluations of dynamic and plant uptake of N in soil amended with 15N-labelled sewage sludge

    International Nuclear Information System (INIS)

    Kchaou, R.; Khelil, M. N.; Rejeb, S.; Gharbi, F.; Henchi, B.; Hernandez, T.; Destain, J. P.

    2010-01-01

    Field experiments were conducted to evaluate the use of a novel 15N isotope technique for comparing the dynamics of N derived from sewage sludge applied to sorghum to the dynamics of N derived from the commercial fertilizer, urea. The treatments included a control, sludge applied at three rates (3, 6 and 9 t/ha, or 113, 226 and 338 kg N/ha) and N-urea applied at three rates (150, 250 and 350 kg N/ha). Recovery of 15N -labelled sludge was similar for the different nitrogen rates applied , with a mean value of 27%. However, the recovery of 15N -urea decreased as the rate of N application increased (from 38% to 27%). Approximately 22% and 19% of the 15N from sludge and urea, respectively, remained in the 0-60 cm layer of soil, most of which was present in the 0-20 cm layer. Furthermore, losses of 15N -labelled fertilizer were not affected by the N fertilization source, and the greatest losses, which were measured in response to the highest N application rate, were 59%. (authors)

  12. Future Applications in Quantitative Isotopic Tracing using Homogeneously Carbon-13 Labelled Plant Material

    International Nuclear Information System (INIS)

    Slaets, Johanna I.F.; Chen, Janet; Resch, Christian; Mayr, Leopold; Weltin, Georg; Heiling, Maria; Gruber, Roman; Dercon, Gerd

    2017-01-01

    Carbon-13 ("1"3C) and nitrogen-15 ("1"5N) labelled plant material is increasingly being used to trace the fate of plant-derived C and N into the atmosphere, soil, water and organisms in many studies, including those investigating the potential of soils to store greenhouse gases belowground. Storage of C in soils can offset and even reduce atmospheric levels of the greenhouse gas, CO_2, and interest in such studies is growing due to problems associated with anthropogenic greenhouse gas emissions impacting climate change. Reduction of N loss in soils is also of great interest, as it reduces release of the greenhouse gas, N_2O, into the atmosphere. However, accurate quantitative tracing of plant-derived C and N in such research is only possible if plant material is labelled both homogeneously and in sufficient quantities.

  13. Method of preparing mercury with an arbitrary isotopic distribution

    Science.gov (United States)

    Grossman, M.W.; George, W.A.

    1986-12-16

    This invention provides for a process for preparing mercury with a predetermined, arbitrary, isotopic distribution. In one embodiment, different isotopic types of Hg[sub 2]Cl[sub 2], corresponding to the predetermined isotopic distribution of Hg desired, are placed in an electrolyte solution of HCl and H[sub 2]O. The resulting mercurous ions are then electrolytically plated onto a cathode wire producing mercury containing the predetermined isotopic distribution. In a similar fashion, Hg with a predetermined isotopic distribution is obtained from different isotopic types of HgO. In this embodiment, the HgO is dissolved in an electrolytic solution of glacial acetic acid and H[sub 2]O. The isotopic specific Hg is then electrolytically plated onto a cathode and then recovered. 1 fig.

  14. Isotope-labeling studies on the formation pathway of acrolein during heat processing of oils.

    Science.gov (United States)

    Ewert, Alice; Granvogl, Michael; Schieberle, Peter

    2014-08-20

    Acrolein (2-propenal) is classified as a foodborne toxicant and was shown to be present in significant amounts in heated edible oils. Up to now, its formation was mainly suggested to be from the glycerol part of triacylglycerides, although a clear influence of the unsaturation of the fatty acid moiety was also obvious in previous studies. To unequivocally clarify the role of the glycerol and the fatty acid parts in acrolein formation, two series of labeled triacylglycerides were synthesized: [(13)C(3)]-triacylglycerides of stearic, oleic, linoleic, and linolenic acid and [(13)C(54)]-triacylglycerides with labeled stearic, oleic, and linoleic acid, but with unlabeled glycerol. Heating of each of the seven intermediates singly in silicon oil and measurement of the formed amounts of labeled and unlabeled acrolein clearly proved the fatty acid backbone as the key precursor structure. Enzymatically synthesized pure linoleic acid and linolenic acid hydroperoxides were shown to be the key intermediates in acrolein formation, thus allowing the discussion of a radical-induced reaction pathway leading to the formation of the aldehyde. Surprisingly, although several oils contained high amounts of acrolein after heating, deep-fried foods themselves, such as donuts or French fries, were low in the aldehyde.

  15. Secretory overexpression and isotopic labeling of the chimeric relaxin family peptide R3/I5 in Pichia pastoris.

    Science.gov (United States)

    Guo, Yu-Qi; Wu, Qing-Ping; Shao, Xiao-Xia; Shen, Ting; Liu, Ya-Li; Xu, Zeng-Guang; Guo, Zhan-Yun

    2015-06-01

    Relaxin family peptides are a group of peptide hormones with divergent biological functions. Mature relaxin family peptides are typically composed of two polypeptide chains with three disulfide linkages, rendering their preparation a challenging task. In the present study, we established an efficient approach for preparation of the chimeric relaxin family peptide R3/I5 through secretory overexpression in Pichia pastoris and in vitro enzymatic maturation. A designed single-chain R3/I5 precursor containing the B-chain of human relaxin-3 and the A-chain of human INSL5 was overexpressed in PichiaPink strain 1 by high-density fermentation in a two-liter fermenter, and approximately 200 mg of purified precursor was obtained from one liter of the fermentation supernatant. We also developed an economical approach for preparation of the uniformly (15)N-labeled R3/I5 precursor by culturing in shaking flasks, and approximately 15 mg of purified (15)N-labeled precursor was obtained from one liter of the culture supernatant. After purification by cation ion-exchange chromatography and reverse-phase high performance liquid chromatography, the R3/I5 precursor was converted to the mature two-chain form by sequential treatment with endoproteinase Lys-C and carboxypeptidase B. The mature R3/I5 peptide had an α-helix-dominated conformation and retained full receptor-binding and receptor activation activities. Thus, Pichia overexpression was an efficient approach for sample preparation and isotopic labeling of the chimeric R3/I5 peptide. This approach could also be extended to the preparation of other relaxin family peptides in future studies.

  16. Expression and isotopic labelling of the potassium channel blocker ShK toxin as a thioredoxin fusion protein in bacteria.

    Science.gov (United States)

    Chang, Shih Chieh; Galea, Charles A; Leung, Eleanor W W; Tajhya, Rajeev B; Beeton, Christine; Pennington, Michael W; Norton, Raymond S

    2012-10-01

    The polypeptide toxin ShK is a potent blocker of Kv1.3 potassium channels, which play a crucial role in the activation of human effector memory T-cells (T(EM)). Selective blockers constitute valuable therapeutic leads for the treatment of autoimmune diseases mediated by T(EM) cells, such as multiple sclerosis, rheumatoid arthritis, and type-1 diabetes. We have established a recombinant peptide expression system in order to generate isotopically-labelled ShK and various ShK analogues for in-depth biophysical and pharmacological studies. ShK was expressed as a thioredoxin fusion protein in Escherichia coli BL21 (DE3) cells and purified initially by Ni²⁺ iminodiacetic acid affinity chromatography. The fusion protein was cleaved with enterokinase and purified to homogeneity by reverse-phase HPLC. NMR spectra of ¹⁵N-labelled ShK were similar to those reported previously for the unlabelled synthetic peptide, confirming that recombinant ShK was correctly folded. Recombinant ShK blocked Kv1.3 channels with a K(d) of 25 pM and inhibited the proliferation of human and rat T lymphocytes with a preference for T(EM) cells, with similar potency to synthetic ShK in all assays. This expression system also enables the efficient production of ¹⁵N-labelled ShK for NMR studies of peptide dynamics and of the interaction of ShK with Kv1.3 channels. Copyright © 2012 Elsevier Ltd. All rights reserved.

  17. Plasma Protein Turnover Rates in Rats Using Stable Isotope Labeling, Global Proteomics, and Activity-Based Protein Profiling

    Energy Technology Data Exchange (ETDEWEB)

    Smith, Jordan N.; Tyrrell, Kimberly J.; Hansen, Joshua R.; Thomas, Dennis G.; Murphree, Taylor A.; Shukla, Anil K.; Luders, Teresa; Madden, James M.; Li, Yunying; Wright, Aaron T.; Piehowski, Paul D.

    2017-12-06

    Protein turnover is important for general health on cellular and organism scales providing a strategy to replace old, damaged, or dysfunctional proteins. Protein turnover also informs of biomarker kinetics, as a better understanding of synthesis and degradation of proteins increases the clinical utility of biomarkers. Here, turnover rates of plasma proteins in rats were measured in vivo using a pulse-chase stable isotope labeling experiment. During the pulse, rats (n=5) were fed 13C6-labeled lysine (“heavy”) feed for 23 days to label proteins. During the chase, feed was changed to an unlabeled equivalent feed (“light”), and blood was repeatedly sampled from rats over 10 time points for 28 days. Plasma samples were digested with trypsin, and analyzed with liquid chromatography-tandem mass spectrometry (LC-MS/MS). MaxQuant was used to identify peptides and proteins, and quantify heavy:light lysine ratios. A system of ordinary differential equations was used to calculate protein turnover rates. Using this approach, 273 proteins were identified, and turnover rates were quantified for 157 plasma proteins with half-lives ranging 0.3-103 days. For the ~70 most abundant proteins, variability in turnover rates among rats was low (median coefficient of variation: 0.09). Activity-based protein profiling was applied to pooled plasma samples to enrich serine hydrolases using a fluorophosphonate (FP2) activity-based probe. This enrichment resulted in turnover rates for an additional 17 proteins. This study is the first to measure global plasma protein turnover rates in rats in vivo, measure variability of protein turnover rates in any animal model, and utilize activity-based protein profiling for enhancing measurements of targeted, low-abundant proteins, such as those commonly used as biomarkers. Measured protein turnover rates will be important for understanding of the role of protein turnover in cellular and organism health as well as increasing the utility of protein

  18. 16 CFR 301.27 - Label and method of affixing.

    Science.gov (United States)

    2010-01-01

    ... all times during the marketing of a fur product the required label shall have a minimum dimension of one and three-fourths (13/4) inches by two and three-fourths (23/4) inches (4.5 cm × 7 cm). Such label...

  19. Sandwich nucleic acid hybridization: a method with a universally usable labeled probe for various specific tests

    International Nuclear Information System (INIS)

    Wolf, H.; Leser, U.; Haus, M.; Gu, S.Y.; Pathmanathan, R.

    1986-01-01

    The use of recombinant m13 phages as hybridization probes offers a considerable advantage over the commonly used recombinant plasmids as the preparation of the DNA probe is very simple and it can easily be labeled directly, e.g. with isotopes with long half-life like 125 I and used for hybridization. However, as the application of nucleic acid hybridization for diagnostic and epidemiological purposes becomes almost unavoidable, the logistic problems of keeping numerous individually labeled hybridization probes increase considerably and may reach prohibitory levels in less well-equipped laboratories. In a new sandwich technique, the first step involves hybridization with an unlabeled recombinant m13 DNA carrying an insert of the desired specificity. In a second step a universally usable labeled probe directed against the m13 part of the recombinant phage DNA is applied. This reduces considerably the problem of preparing and keeping multiple labeled probes in stock. (Auth.)

  20. Determination of bovine lactoferrin in dairy products by ultra-high performance liquid chromatography–tandem mass spectrometry based on tryptic signature peptides employing an isotope-labeled winged peptide as internal standard

    International Nuclear Information System (INIS)

    Zhang, Jingshun; Lai, Shiyun; Cai, Zengxuan; Chen, Qi; Huang, Baifen; Ren, Yiping

    2014-01-01

    Highlights: • A UHPLC–MS/MS method for quantification of bovine lactoferrin was developed. • Tryptic fragment LRPVAAEIYGTK was chosen as signature peptide of bovine lactoferrin. • A winged peptide containing isotopically-labeled signature peptide was designed as internal standard. • The method for determining lactoferrin does not discriminate between the different forms of lactoferrin. • Meet the growing demand to quantify bovine lactoferrin in different dairy products. - Abstract: A new and sensitive determination method was developed for bovine lactoferrin in dairy products including infant formulas based on the signature peptide by ultra high-performance liquid chromatography and triple-quadrupole tandem mass spectrometry under the multiple reaction monitoring mode. The simple pretreatment procedures included the addition of a winged peptide containing the isotope-labeled signature peptide as internal standard, followed by an enzymatic digestion with trypsin. The signature peptide was chosen and identified from the tryptic hydrolyzates of bovine lactoferrin by ultra high-performance liquid chromatography and quadrupole-time-of-flight tandem mass spectrometry based on sequence database search. Analytes were separated on an ACQUITY UPLC BEH 300 C18 column and monitored by MS/MS in seven minutes. Quantitative result bias due to matrix effect and tryptic efficiency was corrected through the use of synthetic isotope-labeled standards. The limit of detection and limit of quantification were 0.3 mg/100 g and 1.0 mg/100 g, respectively. Bovine lactoferrin within the concentration range of 10–1000 nmol L −1 showed a strong linear relationship with a linear correlation coefficient (r) of >0.998. The intra- and inter-day precision of the method were RSD < 6.5% and RSD < 7.1%, respectively. Excellent repeatability (RSD < 6.4%) substantially supported the application of this method for the determination of bovine lactoferrin in dairy samples. The present

  1. Determination of bovine lactoferrin in dairy products by ultra-high performance liquid chromatography–tandem mass spectrometry based on tryptic signature peptides employing an isotope-labeled winged peptide as internal standard

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Jingshun [Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou 310051 (China); Lai, Shiyun [Beingmate Research Institute, Beingmate Baby and Child Food Co., Ltd., Hangzhou 310007 (China); Cai, Zengxuan [Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou 310051 (China); Chen, Qi [Beingmate Research Institute, Beingmate Baby and Child Food Co., Ltd., Hangzhou 310007 (China); Huang, Baifen [Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou 310051 (China); Ren, Yiping, E-mail: renyiping@263.net [Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou 310051 (China)

    2014-06-01

    Highlights: • A UHPLC–MS/MS method for quantification of bovine lactoferrin was developed. • Tryptic fragment LRPVAAEIYGTK was chosen as signature peptide of bovine lactoferrin. • A winged peptide containing isotopically-labeled signature peptide was designed as internal standard. • The method for determining lactoferrin does not discriminate between the different forms of lactoferrin. • Meet the growing demand to quantify bovine lactoferrin in different dairy products. Abstract: A new and sensitive determination method was developed for bovine lactoferrin in dairy products including infant formulas based on the signature peptide by ultra high-performance liquid chromatography and triple-quadrupole tandem mass spectrometry under the multiple reaction monitoring mode. The simple pretreatment procedures included the addition of a winged peptide containing the isotope-labeled signature peptide as internal standard, followed by an enzymatic digestion with trypsin. The signature peptide was chosen and identified from the tryptic hydrolyzates of bovine lactoferrin by ultra high-performance liquid chromatography and quadrupole-time-of-flight tandem mass spectrometry based on sequence database search. Analytes were separated on an ACQUITY UPLC BEH 300 C18 column and monitored by MS/MS in seven minutes. Quantitative result bias due to matrix effect and tryptic efficiency was corrected through the use of synthetic isotope-labeled standards. The limit of detection and limit of quantification were 0.3 mg/100 g and 1.0 mg/100 g, respectively. Bovine lactoferrin within the concentration range of 10–1000 nmol L⁻¹ showed a strong linear relationship with a linear correlation coefficient (r) of >0.998. The intra- and inter-day precision of the method were RSD < 6.5% and RSD < 7.1%, respectively. Excellent repeatability (RSD < 6.4%) substantially supported the application of this method for the determination of bovine lactoferrin in dairy samples. The present method

  2. Competition for phosphorus: differential uptake from dual-isotope-labeled soil interspaces between shrub and grass

    International Nuclear Information System (INIS)

    Caldwell, M.M.; Eissenstat, D.M.; Richards, J.H.; Allen, M.F.

    1985-01-01

    Two species of Agropyron grass differed strikingly in their capacity to compete for phosphate in soil interspaces shared with a common competitor, the sagebrush Artemisia tridentata. Of the total phosphorus-32 and -33 absorbed by Artemisia, 86% was from the interspace shared with Agropyron spicatum and only 14% from that shared with Agropyron desertorum. Actively absorbing mycorrhizal roots of Agropyron and Artemisia were present in both interspaces, where competition for the labeled phosphate occurred. The results have important implications about the way in which plants compete for resources below ground in both natural plant communities and agricultural intercropping systems

  3. System and method for high precision isotope ratio destructive analysis

    Science.gov (United States)

    Bushaw, Bruce A; Anheier, Norman C; Phillips, Jon R

    2013-07-02

    A system and process are disclosed that provide high accuracy and high precision destructive analysis measurements for isotope ratio determination of relative isotope abundance distributions in liquids, solids, and particulate samples. The invention utilizes a collinear probe beam to interrogate a laser ablated plume. This invention provides enhanced single-shot detection sensitivity approaching the femtogram range, and isotope ratios that can be determined at approximately 1% or better precision and accuracy (relative standard deviation).

  4. In Vivo Isotopic Labeling of Symbiotic Bacteria Involved in Cellulose Degradation and Nitrogen Recycling within the Gut of the Forest Cockchafer (Melolontha hippocastani).

    Science.gov (United States)

    Alonso-Pernas, Pol; Bartram, Stefan; Arias-Cordero, Erika M; Novoselov, Alexey L; Halty-deLeon, Lorena; Shao, Yongqi; Boland, Wilhelm

    2017-01-01

    The guts of insects harbor symbiotic bacterial communities. However, due to their complexity, it is challenging to relate a specific symbiotic phylotype to its corresponding function. In the present study, we focused on the forest cockchafer ( Melolontha hippocastani ), a phytophagous insect with a dual life cycle, consisting of a root-feeding larval stage and a leaf-feeding adult stage. By combining in vivo stable isotope probing (SIP) with 13 C cellulose and 15 N urea as trophic links, with Illumina MiSeq (Illumina-SIP), we unraveled bacterial networks processing recalcitrant dietary components and recycling nitrogenous waste. The bacterial communities behind these processes change between larval and adult stages. In 13 C cellulose-fed insects, the bacterial families Lachnospiraceae and Enterobacteriaceae were isotopically labeled in larvae and adults, respectively. In 15 N urea-fed insects, the genera Burkholderia and Parabacteroides were isotopically labeled in larvae and adults, respectively. Additionally, the PICRUSt-predicted metagenome suggested a possible ability to degrade hemicellulose and to produce amino acids of, respectively, 13 C cellulose- and 15 N urea labeled bacteria. The incorporation of 15 N from ingested urea back into the insect body was confirmed, in larvae and adults, by isotope ratio mass spectrometry (IRMS). Besides highlighting key bacterial symbionts of the gut of M. hippocastani , this study provides example on how Illumina-SIP with multiple trophic links can be used to target microorganisms embracing different roles within an environment.

  5. In Vivo Isotopic Labeling of Symbiotic Bacteria Involved in Cellulose Degradation and Nitrogen Recycling within the Gut of the Forest Cockchafer (Melolontha hippocastani

    Directory of Open Access Journals (Sweden)

    Pol Alonso-Pernas

    2017-10-01

    Full Text Available The guts of insects harbor symbiotic bacterial communities. However, due to their complexity, it is challenging to relate a specific symbiotic phylotype to its corresponding function. In the present study, we focused on the forest cockchafer (Melolontha hippocastani, a phytophagous insect with a dual life cycle, consisting of a root-feeding larval stage and a leaf-feeding adult stage. By combining in vivo stable isotope probing (SIP with 13C cellulose and 15N urea as trophic links, with Illumina MiSeq (Illumina-SIP, we unraveled bacterial networks processing recalcitrant dietary components and recycling nitrogenous waste. The bacterial communities behind these processes change between larval and adult stages. In 13C cellulose-fed insects, the bacterial families Lachnospiraceae and Enterobacteriaceae were isotopically labeled in larvae and adults, respectively. In 15N urea-fed insects, the genera Burkholderia and Parabacteroides were isotopically labeled in larvae and adults, respectively. Additionally, the PICRUSt-predicted metagenome suggested a possible ability to degrade hemicellulose and to produce amino acids of, respectively, 13C cellulose- and 15N urea labeled bacteria. The incorporation of 15N from ingested urea back into the insect body was confirmed, in larvae and adults, by isotope ratio mass spectrometry (IRMS. Besides highlighting key bacterial symbionts of the gut of M. hippocastani, this study provides example on how Illumina-SIP with multiple trophic links can be used to target microorganisms embracing different roles within an environment.

  6. A microautoradiographic method permitting the study of 99m-technetium labelled leukocytes

    International Nuclear Information System (INIS)

    Colas-Linhart, N.; Perianin, A.; Petiet, A.; Bretillon, A.; Bok, B.

    1985-01-01

    Cell migration studies are very important in inflammatory phenomena. Methods currently used are not quantitative and have been subject to much controversy. Homogeneity of sup(99m)Tc leukocyte labelling was verified by a microautoradiographic method (MAR), which was performed in our laboratory. This method was used in cell migration studies to verify if the migrating cells were indeed the labelled cells [fr

  7. Calculation of isotopic mass and energy production by a matrix operator method

    International Nuclear Information System (INIS)

    Lee, C.E.

    1976-08-01

    The Volterra method of the multiplicative integral is used to determine the isotopic density, mass, and energy production in linear systems. The solution method, assumptions, and limitations are discussed. The method allows a rapid accurate calculation of the change in isotopic density, mass, and energy production independent of the magnitude of the time steps, production or decay rates, or flux levels

  8. Easy Extraction Method To Evaluate δ13C Vanillin by Liquid Chromatography-Isotopic Ratio Mass Spectrometry in Chocolate Bars and Chocolate Snack Foods.

    Science.gov (United States)

    Bononi, Monica; Quaglia, Giancarlo; Tateo, Fernando

    2015-05-20

    An easy extraction method that permits the use of a liquid chromatography-isotopic ratio mass spectrometry (LC-IRMS) system to evaluate δ(13)C of vanillin in chocolate products and industrial flavorings is presented. The method applies the determination of stable isotopes of carbon to discriminate between natural vanillin from vanilla beans and vanillin from other sources (mixtures from beans, synthesis, or biotechnology). A series of 13 chocolate bars and chocolate snack foods available on the Italian market and 8 vanilla flavorings derived from industrial quality control processes were analyzed. Only 30% of products considered in this work that declared "vanilla" on the label showed data that permitted the declaration "vanilla" according to European Union (EU) Regulation 1334/2008. All samples not citing "vanilla" or "natural flavoring" on the label gave the correct declaration. The extraction method is presented with data useful for statistical evaluation.

  9. Evaluation of isotopic dilution method for measuring N2 fixation in azolla: comparison with other methods

    International Nuclear Information System (INIS)

    Sah, R.N.; Goyal, S.S.; Rains, D.W.; Paige, D.F.

    1989-01-01

    An isotopic dilution method that overcomes the drawbacks of commonly used methods for measuring N 2 fixation by aquatic N‐fixers such as Azolla pinnata‐Anabaena azollae association (Azolla) is presented. The method was compared with 15 N2 gas (while maintaining CO 2 ) and the difference methods of measuring N 2 fixation. The isotopic dilution method was used for two conditions: a. For 15 N‐free growth medium, Azolla was pre‐enriched with 15 N, and N 2 fixation was determined by measuring the dilution of 15 N in the tissue. b. For the growth medium containing N, N2 fixation was determined by providing 15 N enriched ammonium sulfate in the growth medium and measuring 15 N to 14 N ratio in the tissue. An airtight chamber, necessary for 15 N 2 gas and acetylene reduction methods, was not representative of the growing environment of Azolla. Temperature in the airtight chamber was far from uniform and CO 2 was rapidly depleted. The isotopic dilution method is simpler, relatively inexpensive, subject to fewer errors and applicable to more diverse conditions, and yet was as accurate as 15 N2‐gas method. (author)

  10. Dual isotope assays

    International Nuclear Information System (INIS)

    Smith, G.F.W.; Stevens, R.A.J.; Jacoby, B.

    1980-01-01

    Dual isotope assays for thyroid function are performed by carrying out a radio-immunoassay for two of thyroxine (T4), tri-iodothyronine (T3), thyroid stimulating hormone (TSH), and thyroxine binding globulin (TBG), by a method wherein a version of one of the thyroid components, preferably T4 or T3 is labelled with Selenium-75 and the version of the other thyroid component is labelled with a different radionuclide, preferably Iodine-125. (author)

  11. GC-MS DETERMINATION OF RATIOS OF STABLE-ISOTOPE LABELED TO NATURAL UREA USING [(CN2)-C-13-N-15]UREA FOR STUDYING UREA KINETICS IN SERUM AND AS A MEANS TO VALIDATE ROUTINE METHODS FOR THE QUANTITATIVE ASSAY OF UREA IN DIALYSATE

    NARCIS (Netherlands)

    WOLTHERS, BG; TEPPER, T; WITHAG, A; NAGEL, GT; DEHAAN, THY; VANLEEUWEN, JJ; STEGEMAN, CA; HUISMAN, RM

    A GC-MS determination of urea in serum or spent dialysate is described, using (CN2)-C-13-N-15-labelled urea and assaying the area ratio of labelled to natural urea by mass fragmentographic monitoring of fragments m/e 153 and 156, after its eventual conversion into the trimethylsilylether-derivative

  12. Utilization of carbon 13-labelled stable isotopes for studying drug toxicity on cellular metabolism

    International Nuclear Information System (INIS)

    Herve, M.; Wietzerbin, J.; Tran-Dinh, S.

    1994-01-01

    A new approach for studying the effects of two drugs, amphotericine B (AMB), an anti-fungal antibiotic, and 2-deoxy-D-glucose (DG), on the glucose metabolism in brewer yeast cells (Saccharomyces cerevisiae), is presented; AMB interacts with the membrane sterols, inducing formation of pores through which ions and small molecules can pass. DG may enter in the cytosol, where it is phosphoryled by hexokinase into deoxy-D-glucose 6-phosphate (DG6P) which disappears very slowly. DG slows down the glycolysis process and induces the formation of new substances. This paper shows the advantages of utilizing carbon 13-labelled substrates combined to the NMR-13C and NMR-1H techniques. 6 figs., 5 refs

  13. Understanding N timing in corn yield and fertilizer N recovery: An insight from an isotopic labeled-N determination

    Science.gov (United States)

    de Almeida, Rodrigo Estevam Munhoz; Pierozan Junior, Clovis; Lago, Bruno Cocco; Trivelin, Paulo Cesar Ocheuze

    2018-01-01

    Early fertilizer nitrogen (N) application on cover crops or their residues during the off-season is a practice adopted in Brazil subtropical conditions under no-tillage corn (Zea mays L.) systems. However, the effect of early N application on yield, plant N content, and N recovery efficiency (NRE) for corn is not yet well documented. Five fertilizer N timings in an oat-corn system were evaluated in two studies utilizing an isotopic-labeled N determination, 15N isotope. The N fertilization timings were: (i) oat tillering, (ii) 15 days before corn planting time, over the oat residues, (iii) at corn planting time, (iv) in-season at the three-leaf growth stage (V3), and (v) in-season split application at V3 and six-leaf (V6) growth stages. Based on the statistical analysis, the N fertilization timings were separated into three groups: 1) N-OATS, designated to N applied at oat; 2) N-PLANT, referred to pre-plant and planting N applications; and 3) N-CORN, designated to in-season corn N applications. Corn yield was not affected by the N fertilization timing. However, the N-CORN N fertilization timings enhanced NRE by 17% and 35% and final N recovery system (plant plus soil) by 16% and 24% all relative to N-OATS and N-PLANT groups, respectively. Overall, N-OATS resulted in the largest N derived from fertilizer (NDFF) amount in the deeper soil layer, in overall a delta of 10 kg N ha-1 relative to the rest of the groups. Notwithstanding corn yield was not affected, early N fertilization under subtropical conditions is not a viable option since NRE was diminished and the non-recovery N increased relative to the in-season N applications. PMID:29462178

  14. Understanding N timing in corn yield and fertilizer N recovery: An insight from an isotopic labeled-N determination.

    Science.gov (United States)

    Maciel de Oliveira, Silas; Almeida, Rodrigo Estevam Munhoz de; Ciampitti, Ignacio A; Pierozan Junior, Clovis; Lago, Bruno Cocco; Trivelin, Paulo Cesar Ocheuze; Favarin, José Laércio

    2018-01-01

    Early fertilizer nitrogen (N) application on cover crops or their residues during the off-season is a practice adopted in Brazil subtropical conditions under no-tillage corn (Zea mays L.) systems. However, the effect of early N application on yield, plant N content, and N recovery efficiency (NRE) for corn is not yet well documented. Five fertilizer N timings in an oat-corn system were evaluated in two studies utilizing an isotopic-labeled N determination, 15N isotope. The N fertilization timings were: (i) oat tillering, (ii) 15 days before corn planting time, over the oat residues, (iii) at corn planting time, (iv) in-season at the three-leaf growth stage (V3), and (v) in-season split application at V3 and six-leaf (V6) growth stages. Based on the statistical analysis, the N fertilization timings were separated into three groups: 1) N-OATS, designated to N applied at oat; 2) N-PLANT, referred to pre-plant and planting N applications; and 3) N-CORN, designated to in-season corn N applications. Corn yield was not affected by the N fertilization timing. However, the N-CORN N fertilization timings enhanced NRE by 17% and 35% and final N recovery system (plant plus soil) by 16% and 24% all relative to N-OATS and N-PLANT groups, respectively. Overall, N-OATS resulted in the largest N derived from fertilizer (NDFF) amount in the deeper soil layer, in overall a delta of 10 kg N ha-1 relative to the rest of the groups. Notwithstanding corn yield was not affected, early N fertilization under subtropical conditions is not a viable option since NRE was diminished and the non-recovery N increased relative to the in-season N applications.

  15. Quantifying the effect of plant growth on litter decomposition using a novel, triple-isotope label approach

    Science.gov (United States)

    Ernakovich, J. G.; Baldock, J.; Carter, T.; Davis, R. A.; Kalbitz, K.; Sanderman, J.; Farrell, M.

    2017-12-01

    Microbial degradation of plant detritus is now accepted as a major stabilizing process of organic matter in soils. Most of our understanding of the dynamics of decomposition come from laboratory litter decay studies in the absence of plants, despite the fact that litter decays in the presence of plants in many native and managed systems. There is growing evidence that living plants significantly impact the degradation and stabilization of litter carbon (C) due to changes in the chemical and physical nature of soils in the rhizosphere. For example, mechanistic studies have observed stimulatory effects of root exudates on litter decomposition, and greenhouse studies have shown that living plants accelerate detrital decay. Despite this, we lack a quantitative understanding of the contribution of living plants to litter decomposition and how interactions of these two sources of C build soil organic matter (SOM). We used a novel triple-isotope approach to determine the effect of living plants on litter decomposition and C cycling. In the first stage of the experiment, we grew a temperate grass commonly used for forage, Poa labillardieri, in a continuously-labelled atmosphere of 14CO2 fertilized with K15NO3, such that the grass biomass was uniformly labelled with 14C and 15N. In the second stage, we constructed litter decomposition mescososms with and without a living plant to test for the effect of a growing plant on litter decomposition. The 14C/15N litter was decomposed in a sandy clay loam while a temperate forage grass, Lolium perenne, grew in an atmosphere of enriched 13CO2. The fate of the litter-14C/15N and plant-13C was traced into soil mineral fractions and dissolved organic matter (DOM) over the course of nine weeks using four destructive harvests of the mesocosms. Our preliminary results suggest that living plants play a major role in the degradation of plant litter, as litter decomposition was greater, both in rate and absolute amount, for soil mesocosms

  16. Method for enriching and separating heavy hydrogen isotopes from substance streams containing such isotopes by means of isotope exchange

    International Nuclear Information System (INIS)

    Knochel, A.; Eggers, I.; Klatte, B.; Wilken, R. D.

    1985-01-01

    A process for enriching and separating heavy hydrogen isotopes having a heavy hydrogen cation (deuterium and/or tritium) from substance streams containing them, wherein the respectively present hydrogen isotopes are exchanged in chemical equilibria. A protic, acid solution containing deuterium and/or tritium is brought into contact with a value material from the group of open-chained polyethers or aminopolyethers, macro-monocyclic or macro-polycyclic polyethers, macro-monocyclic or macro-polycyclic amino polyethers, and mixtures of these values, in their free or proton salt form to form a reaction product of the heavy hydrogen cation with the value or value salt and bring about enrichment of deuterium and/or tritium in the reaction product. The reaction product containing the value or value salt is separated from the solution. The separated reaction product is treated to release the hydrogen isotope(s) to be enriched in the form of deuterium oxide (HDO) and/or tritium oxide (HTO) by regenerating the value or its salt, respectively. The regenerated value is returned for reuse

  17. Labelling of blood cells with radioactive indium-201: method, results, indications

    International Nuclear Information System (INIS)

    Ducassou, D.; Brendel, A.; Nouel, J.P.

    1978-01-01

    A modification of the method of Thakur et al. for labelling polynuclear cells with 8-hydroxyquinolein-indium-complexe utilising the water soluble sulfate of the substance was applied. The labelling procedure gave a yield over 98% with erthrocytes and over 80% with platelets and polynuclear cells using at least 1 x 10 8 plasma free cells. Functional capacity of the labelled cells remained unaltered. Injection double labelled ( 111 In, 51 Cr) red cells correlation of values for the red cell volume amounted to r = 0,98 (n=20); red cell life-spane measurements gave comparable results in 5 patients. After injecting labelled platelets a life-spane between 6,5 and 11 days was measured. Scintigraphic visualisation of pulmonary embolism was obtained 30 minutes after injecting labelled platelets. Injection of labelled polynuclear cells allows life-spane measurements as well as detection of abscesses. (author)

  18. The fluorodediazonation - a method for n.c.a.-18F-labelling of aromatic substrates

    International Nuclear Information System (INIS)

    Zwernemann, O.

    1991-06-01

    For the positron emission tomography (PET) applications, radiopharmaceuticals are required that are labelled with short-lived positron emitters. Fluorine-18 has become the leading radionuclide used for PET, due to its favourable physical properties. However, the labelling of aromatic substances with fluorine-18 with the methods available presents problems not encountered with aliphatic compounds. The decomposition of aromatic diazonium salts opens up feasible ways of preparing a broad range of labelled compounds. The dissertation investigated the possibilities of labelling with fluorine-18 by way of dediazonation on the standard substrate p-Toluidyl diazonium ion. The results reported show that the method of fluorodediazonation is an interesting further method for F-18 labelling of aromatic substrates in addition to the hitherto applied techniques. It allows carrier-free labelling of a large group of substances which cannot be fluorinated via direct nucleophilicity. (BBR) [de

  19. Auto-inducing media for uniform isotope labeling of proteins with {sup 15}N, {sup 13}C and {sup 2}H

    Energy Technology Data Exchange (ETDEWEB)

    Guthertz, Nicolas [Institute of Cancer Research, Division of Structural Biology (United Kingdom); Klopp, Julia; Winterhalter, Aurélie; Fernández, César; Gossert, Alvar D., E-mail: alvar.gossert@novartis.com [Novartis Institutes for BioMedical Research (Switzerland)

    2015-06-15

    Auto-inducing media for protein expression offer many advantages like robust reproducibility, high yields of soluble protein and much reduced workload. Here, an auto-inducing medium for uniform isotope labelling of proteins with {sup 15}N, {sup 13}C and/or {sup 2}H in E. coli is presented. So far, auto-inducing media have not found widespread application in the NMR field, because of the prohibitively high cost of labeled lactose, which is an essential ingredient of such media. Here, we propose using lactose that is only selectively labeled on the glucose moiety. It can be synthesized from inexpensive and readily available substrates: labeled glucose and unlabeled activated galactose. With this approach, uniformly isotope labeled proteins were expressed in unattended auto-inducing cultures with incorporation of {sup 13}C, {sup 15}N of 96.6 % and {sup 2}H, {sup 15}N of 98.8 %. With the present protocol, the NMR community could profit from the many advantages that auto-inducing media offer.

  20. An innovative method for extracting isotopic information from low-resolution gamma spectra

    International Nuclear Information System (INIS)

    Miko, D.; Estep, R.J.; Rawool-Sullivan, M.W.

    1998-01-01

    A method is described for the extraction of isotopic information from attenuated gamma ray spectra using the gross-count material basis set (GC-MBS) model. This method solves for the isotopic composition of an unknown mixture of isotopes attenuated through an absorber of unknown material. For binary isotopic combinations the problem is nonlinear in only one variable and is easily solved using standard line optimization techniques. Results are presented for NaI spectrum analyses of various binary combinations of enriched uranium, depleted uranium, low burnup Pu, 137 Cs, and 133 Ba attenuated through a suite of absorbers ranging in Z from polyethylene through lead. The GC-MBS method results are compared to those computed using ordinary response function fitting and with a simple net peak area method. The GC-MBS method was found to be significantly more accurate than the other methods over the range of absorbers and isotopic blends studied

  1. Radiation Oxidation Mechanisms in Polyolefins Studied by C-13 Isotopic Labeling

    International Nuclear Information System (INIS)

    Clough, R.L.

    2006-01-01

    Control of oxidative degradation is a critical consideration in most applications involving polymers and radiation. In radiation crosslinking or sterilization, or in the use of polymers in radiation environments (such as nuclear plants), the objective is to minimize degradation as much as possible. In other applications, a controlled, partial degradation is desired to alter processing properties, or to enhance adhesion or solubility. To gain more understanding of the complex processes of radiation oxidation, samples of one important commercial polyolefin, polypropylene, were synthesized in which the three different carbon atoms along the chain were selectively labeled with carbon-13. These samples were subjected to radiation under inert and air atmospheres, and to post-irradiation thermal exposure in air at various temperatures. Analysis of macromolecular radiation-oxidation products was carried out using 13 C NMR and FTIR. Time-dependent plots of oxidation products have been obtained from the NMR measurements, including the post-irradiation oxidation of a sample held at room temperature in air that has been monitored for 2 years. Analysis of volatile oxidation products (CO, CO 2 , and small organic molecules) was accomplished with gas chromatography / mass spectroscopy. The position of the 13 C labels in the degradation products, have been traced back to their positions of origin on the macromolecule, providing insights into the chemical reaction mechanisms through which the products were formed. The major solid-phase products include peroxides and alcohols, both formed at tertiary carbon sites along the chain. Other products include methyl ketones, acids, esters, peresters, and hemiketals formed from reaction at the tertiary carbon, together with in-chain ketones and esters from reaction at the secondary chain carbon. No evidence is found of macromolecular products arising from reactions at the methyl side chain. Significant temperature-dependent differences are

  2. Statistical methods for quantitative mass spectrometry proteomic experiments with labeling

    Directory of Open Access Journals (Sweden)

    Oberg Ann L

    2012-11-01

    Full Text Available Abstract Mass Spectrometry utilizing labeling allows multiple specimens to be subjected to mass spectrometry simultaneously. As a result, between-experiment variability is reduced. Here we describe use of fundamental concepts of statistical experimental design in the labeling framework in order to minimize variability and avoid biases. We demonstrate how to export data in the format that is most efficient for statistical analysis. We demonstrate how to assess the need for normalization, perform normalization, and check whether it worked. We describe how to build a model explaining the observed values and test for differential protein abundance along with descriptive statistics and measures of reliability of the findings. Concepts are illustrated through the use of three case studies utilizing the iTRAQ 4-plex labeling protocol.

  3. Statistical methods for quantitative mass spectrometry proteomic experiments with labeling.

    Science.gov (United States)

    Oberg, Ann L; Mahoney, Douglas W

    2012-01-01

    Mass Spectrometry utilizing labeling allows multiple specimens to be subjected to mass spectrometry simultaneously. As a result, between-experiment variability is reduced. Here we describe use of fundamental concepts of statistical experimental design in the labeling framework in order to minimize variability and avoid biases. We demonstrate how to export data in the format that is most efficient for statistical analysis. We demonstrate how to assess the need for normalization, perform normalization, and check whether it worked. We describe how to build a model explaining the observed values and test for differential protein abundance along with descriptive statistics and measures of reliability of the findings. Concepts are illustrated through the use of three case studies utilizing the iTRAQ 4-plex labeling protocol.

  4. Method for adding additional isotopes to actinide-only burnup credit

    International Nuclear Information System (INIS)

    Lancaster, D.B.; Fuentes, E.; Kang, C.

    1998-01-01

    The Topical Report on Actinide-Only Burnup Credit for Pressurized Water Reactor Spent Nuclear Fuel Packages requires computer code validation to be performed against a benchmark set of chemical assays for isotopic concentration and against a benchmark set of critical experiments for package criticality. Both sets contain all the isotopes included in the methodology. The chemical assays used include the uranium and plutonium isotopes, while the critical experiments were composed of UO 2 or MOX rods, covering the isotopes in the actinide only approach. Since other isotopes are not included in the validation benchmark sets, it would be necessary to justify both the content and worth of any additional isotope for which burnup credit is to be taken (i.e., both the concentration and criticality effect of each particular isotope must be validated). A method is proposed here that can be used for any number of additional isotopes. As does the actinide-only burnup credit methodology, this method makes use of chemical assay data to establish the conservatism in the prediction of each isotope's concentration. Criticality validation is also performed using a benchmark set of UO 2 and MOX critical experiments, where the additional isotopes are validated using worth experiments to conservatively account for any uncertainty in their cross sections. The remaining requirements (analysis and modeling parameters, loading criteria generation, and physical implementation and controls) are performed exactly as described in the actinide-only burnup credit methodology. This report provides insight into each particular requirement in the new methodology

  5. Methods for the synthesis of tritium labelled steroids

    International Nuclear Information System (INIS)

    Volkova, V.S.; Tatarkina, F.V.; Kaklyushkina, L.N.; Ignat'eva, N.A.; Tupitsyn, I.F.; Efimova, Ts.I.

    1977-01-01

    The catalytic substitution of bromine for tritium in corresponding bromo-derivatives of steroids was used for the preparation of 4 steroids labelled with tritium at position 7. The bromination of the starting steroids was effected with N-bromosuccinimide. Ten steroids labelled with tritium at the positions 1, 2, 6 and 7 were prepared by reduction of the unsaturated derivatives with gaseous tritium in the presence of either the heterogeneous Pd/C catalyst, or the homogeneous chloride of tris(triphenylphosphine)rhodium

  6. Measurement of loss rates of organic compounds in snow using in situ experiments and isotopically labelled compounds

    Directory of Open Access Journals (Sweden)

    Erika von Schneidemesser

    2012-07-01

    Full Text Available Organic molecular marker compounds are widely used to identify emissions from anthropogenic and biogenic air pollution sources in atmospheric samples and in deposition. Specific organic compounds have been detected in polar regions, but their fate after deposition to snow is poorly characterized. Within this context, a series of exposure experiments were carried out to observe the post-depositional processing of organic compounds under real-world conditions in snow on the surface of the Greenland Ice Sheet, at the Summit research station. Snow was prepared from water spiked with isotopically labelled organic compounds, representative of typical molecular marker compounds emitted from anthropogenic activities. Reaction rate constants and reaction order were determined based on a decrease in concentration to a stable, non-zero, threshold concentration. Fluoranthene-d10, docosane-d46, hexadecanoic acid-d31, docosanoic acid-d43 and azelaic acid-d14 were estimated to have first order loss rates within surface snow with reaction rate constants of 0.068, 0.040, 0.070, 0.067 and 0.047 h−1, respectively. No loss of heptadecane-d36 was observed. Overall, these results suggest that organic contaminants are archived in polar snow, although significant post-depositional losses of specific organic compounds occur. This has implications for the environmental fate of organic contaminants, as well as for ice-core studies that seek to use organic molecular markers to infer past atmospheric loadings, and source emissions.

  7. Production, purification and detergent exchange of isotopically labeled Bacillussubtilis cytochrome b₅₅₈ (SdhC).

    Science.gov (United States)

    Baureder, Michael; Hederstedt, Lars

    2011-11-01

    Cytochrome b₅₅₈ of the gram-positive bacterium Bacillussubtilis is the membrane anchor subunit of the succinate:quinone oxidoreductase of the citric acid cycle. The cytochrome consists of the SdhC polypeptide (202 residues) and two protoheme IX groups that function in transmembrane electron transfer to menaquinone. The general structure of the cytochrome is known from extensive experimental studies and by comparison to Wolinellasuccinogenes fumarate reductase for which the X-ray crystal structure has been determined. Solution state NMR can potentially be used to identify the quinone binding site(s) and study, e.g. redox-linked, dynamics of cytochrome b₅₅₈. In this work we present an efficient procedure for the isolation of preparative amounts of isotopically labeled B. subtilis cytochrome b₅₅₈ produced in Escherichia coli. We have also evaluated several detergents suitable for NMR for their effectiveness in maintaining the cytochrome solubilized and intact for days at room temperature. Copyright © 2011 Elsevier Inc. All rights reserved.

  8. Preparation of radiopharmaceuticals labelled with bromine positron emitting isotopes for the study of dopaminergic receptors of the central nervous system using positron emission tomography

    International Nuclear Information System (INIS)

    Loc'h, C.

    1988-04-01

    The in vivo study of dopaminergic receptors of the central nervous system using positron emission tomography requires the preparation of radiopharmaceuticals labelled with β + emitting isotopes. The chemical and pharmacological properties of these ligands are evaluated. Cyclotron produced 75 and 76 bromine β + emitting isotopes are incorporated into dopaminergic ligands by electrophilic substitution using peracetic acid in a no-carrier added form. Purity, lipophilicity and specific activity are analyzed. Pharmacological criteria (specificity, saturability, displacement, localization) required for ligand-receptor binding studies are evaluated in vitro on striatal membranes and in vivo in the rat. Positron emission tomographic studies show that the study of dopaminergic D2 receptors is possible using 75 and 76 bromine labelled bromospiperone and bromolisuride. These ligands are used in physiological and pharmacological studies of the central nervous system [fr

  9. Isotope methods for the control of food products and beverages

    Energy Technology Data Exchange (ETDEWEB)

    Guillou, C; Reniero, F [Commission of the European Communities, Joint Research Centre, Ispra (Italy)

    2001-10-01

    The measurement of the stable isotope contents provides useful information for the detection of many frauds in food products. Nuclear magnetic resonance (NMR) and isotopic ratio mass spectroscopy (IRMS) are the two main analytical techniques used for the determination of stable isotope contents in food products. These analytical techniques have been considerably improved in the last years offering wider possibilities of applications for food analysis. A review of the applications for the control of food products and beverages is presented. The need for new reference materials is discussed. (author)

  10. Isotope methods for the control of food products and beverages

    International Nuclear Information System (INIS)

    Guillou, C.; Reniero, F.

    2001-01-01

    The measurement of the stable isotope contents provides useful information for the detection of many frauds in food products. Nuclear magnetic resonance (NMR) and isotopic ratio mass spectroscopy (IRMS) are the two main analytical techniques used for the determination of stable isotope contents in food products. These analytical techniques have been considerably improved in the last years offering wider possibilities of applications for food analysis. A review of the applications for the control of food products and beverages is presented. The need for new reference materials is discussed. (author)

  11. Molecularly imprinted solid phase extraction using stable isotope labeled compounds as template and liquid chromatography-mass spectrometry for trace analysis of bisphenol A in water sample

    International Nuclear Information System (INIS)

    Kawaguchi, Migaku; Hayatsu, Yoshio; Nakata, Hisao; Ishii, Yumiko; Ito, Rie; Saito, Koichi; Nakazawa, Hiroyuki

    2005-01-01

    We have developed a molecularly imprinted polymer (MIP) using a stable isotope labeled compound as the template molecule and called it the ''isotope molecularly imprinted polymer'' (IMIP). In this study, bisphenol A (BPA) was used as the model compound. None imprinted polymer (NIP), MIP, dummy molecularly imprinted polymer (DMIP) and IMIP were prepared by the suspension polymerization method using without template, BPA, 4-tert-butylphenol (BP) and bisphenol A-d 16 (BPA-d 16 ), respectively. The polymers were subjected to molecularly imprinted solid phase extraction (MI-SPE), and the extracted samples were subjected to liquid chromatography-mass spectrometry (LC-MS). Although the leakage of BPA-d 16 from the IMIP was observed and that of BPA was not observed. The selectivity factors of MIP and IMIP for BPA were 4.45 and 4.43, respectively. Therefore, IMIP had the same molecular recognition ability as MIP. When MI-SPE with IMIP was used and followed by LC-MS in the analysis of river water sample, the detection limit of BPA was 1 ppt with high sensitivity. Moreover, the average recovery was higher than 99.8% (R.S.D.: 3.7%) by using bisphenol A- 13 C 12 (BPA- 13 C 12 ) as the surrogate standard. In addition, the IMIP were employed in MI-SPE of BPA in river water sample by LC-MS. The concentration of BPA in the river water sample was determined to be 32 pg ml -1 . We confirmed that it was possible to measure trace amounts of a target analyte by MI-SPE using IMIP

  12. Study on atmospheric hydrogen enrichment by cryopump method and isotope separation by gas chromatography

    International Nuclear Information System (INIS)

    Taniyama, Yuki; Momoshima, Noriyuki

    2001-01-01

    To obtain the information of source of atmospheric hydrogen tritium an analysis of tritium isotopes is thought to be effective. So an atmospheric hydrogen enrichment apparatus and a cryogenic gas chromatographic column were made. Experiments were carried out to study the performance of cryopump to enrich atmospheric hydrogen and the column to separate hydrogen isotopes that obtained by cryopump method. The cryopump was able to process about 1000 1 atmosphere and the column was able to separate hydrogen isotopes with good resolution. (author)

  13. Development, optimisation, and application of ICP-SFMS methods for the measurement of isotope ratios

    International Nuclear Information System (INIS)

    Stuerup, S.

    2000-07-01

    The measurement of isotopic composition and isotope ratios in biological and environmental samples requires sensitive, precise, and accurate analytical techniques. The analytical techniques used are traditionally based on mass spectrometry, among these techniques is the ICP-SFMS technique, which became commercially available in the mid 1990s. This technique is characterised by high sensitivity, low background, and the ability to separate analyte signals from spectral interferences. These features are beneficial for the measurement of isotope ratios and enable the measurement of isotope ratios of elements, which it has not previously been possible to measure due to either spectral interferences or poor sensitivity. The overall purpose of the project was to investigate the potential of the single detector ICP-SFMS technique for the measurement of isotope ratios in biological and environmental samples. One part of the work has focused on the fundamental aspects of the ICP-SFMS technique with special emphasize on the features important to the measurement of isotope ratios, while another part has focused on the development, optimisation and application of specific methods for the measurement of isotope ratios of elements of nutritional interest and radionuclides. The fundamental aspects of the ICP-SFMS technique were investigated theoretically and experimentally by the measurement of isotope ratios applying different experimental conditions. It was demonstrated that isotope ratios could be measured reliably using ICP-SFMS by educated choice of acquisition parameters, scanning mode, mass discrimination correction, and by eliminating the influence of detector dead time. Applying the knowledge gained through the fundamental study, ICP-SFMS methods for the measurement of isotope ratios of calcium, zinc, molybdenum and iron in human samples and a method for the measurement of plutonium isotope ratios and ultratrace levels of plutonium and neptunium in environmental samples

  14. Stable isotope labeling confirms mixotrophic nature of streamer biofilm communities at alkaline hot springs

    Directory of Open Access Journals (Sweden)

    Florence eSchubotz

    2015-02-01

    Full Text Available Streamer biofilm communities (SBC are often observed within chemosynthetic zones of Yellowstone hot spring outflow channels, where temperatures exceed those conducive to photosynthesis. Nearest the hydrothermal source (75-88°C SBC comprise thermophilic Archaea and Bacteria, often mixed communities including Desulfurococcales and uncultured Crenarchaeota, as well as Aquificae, Thermus, each carrying diagnostic membrane lipid biomarkers. We tested the hypothesis that SBC can alternate their metabolism between autotrophy and heterotrophy depending on substrate availability. Feeding experiments were performed at two alkaline hot springs in Yellowstone National Park: Octopus Spring and ‘Bison Pool’, using various 13C-labeled substrates (bicarbonate, formate, acetate and glucose to determine the relative uptake of these different carbon sources. Highest 13C uptake, at both sites, was from acetate into almost all bacterial fatty acids, particularly into methyl-branched C15, C17 and C19 fatty acids that are diagnostic for Thermus/Meiothermus and some Firmicutes as well as into universally common C16:0 and C18:0 fatty acids. 13C-glucose showed a similar, but a 10 to 30 times lower uptake across most fatty acids. 13C bicarbonate uptake, signifying the presence of autotrophic communities was only significant at ‘Bison Pool’ and was observed predominantly in non-specific saturated C16, C18, C20 and C22 fatty acids. Incorporation of 13C-formate occurred only at very low rates at ‘Bison Pool’ and was almost undetectable at Octopus Spring, suggesting that formate is not an important carbon source for SBC. 13C uptake into archaeal lipids occurred predominantly with 13C acetate, suggesting also that archaeal communities at both springs have primarily heterotrophic carbon assimilation pathways. We hypothesize that these communities are energy-limited and predominantly nurtured by input of exogenous organic material, with only a small fraction being

  15. Fatty acids labelled with iodine 123 or 131 in. omega. position; myocardial evolution

    Energy Technology Data Exchange (ETDEWEB)

    Riche, F.; Vidal, M. (Grenoble-1 Univ., 38 (France)); Mathieu, J.P.; Busquet, G.; Comet, M. (Grenoble-1 Univ., 38 - La Tronche (France)); Coornaert, S.; Bardy, A. (CEA Centre d' Etudes Nucleaires de Saclay, 91 - Gif-sur-Yvette (France). Office des Rayonnements Ionisants); Godart, J. (Grenoble-1 Univ., 38 (France). Inst. des Sciences Nucleaires)

    A simple and rapid method of labelling a number of saturated acetylenic and Z or E ethylenic acids has been developed. The fatty acids are labelled with /sup 123/I- or /sup 131/I- in the ..omega.. position by isotopic exchange labelled NaI in acetone. Myocardial metabolism was studied by injecting the labelled fatty acids into mice.

  16. Optimization of in vitro cell labeling methods for human umbilical cord-derived mesenchymal stem cells.

    Science.gov (United States)

    Tao, R; Sun, T-J; Han, Y-Q; Xu, G; Liu, J; Han, Y-F

    2014-01-01

    Human umbilical cord-derived mesenchymal stem cells (hUCMSCs) are a novel source of seed cells for cell therapy and tissue engineering. However, in vitro labeling methods for hUCMSCs need to be optimized for better detection of transplanted cells. To identify the most stable and efficient method for labeling hUCMSCs in vitro. hUCMSCs were isolated using a modified enzymatic digestion procedure and cultured. hUCMSCs of passage three (P3) were then labeled with BrdU, PKH26, or lentivirus-GFP and passaged further. Cells from the first labeled passage (LP1), the fourth labeled passage (LP4) and later passages were observed using a fluorescence microscope. The differentiation potential of LP4 cells was assessed by induction with adipogenic and osteogenic medium. Flow cytometry was used to measure the percentage of labeled cells and the percentage of apoptotic or dead cells. The labeling efficiencies of the three hUCMSC-labeling methods were compared in vitro. BrdU, PKH26, and lentivirus-GFP all labeled LP1 cells with high intensity and clarity. However, the BrdU labeling of the LP4 cells was vague and not localized to the cell nuclei; LP9 cells were not detected under a fluorescence microscope. There was also a significant decrease in the fluorescence intensity of PKH26-labeled LP4 cells, and LP11 cells were not detected under a fluorescence microscope. However, the fluorescence of LP4 cells labeled with lentivirus-GFP remained strong, and cells labeled with lentivirus-GFP were detected up to LP14 under a fluorescence microscope. Statistical analyses indicated that percentages of LP1 cells labeled with PKH26 and lentivirus-GFP were significantly higher than that of cells labeled with BrdU (p 0.05) was observed between the death rates of labeled and unlabeled cells. Lentivirus-GFP is a valid method for long-term in vitro labeling, and it may be used as a long-term hUCMSC tracker following transplantation in vivo.

  17. Analytical performance of reciprocal isotope labeling of proteome digests for quantitative proteomics and its application for comparative studies of aerobic and anaerobic Escherichia coli proteomes

    International Nuclear Information System (INIS)

    Lo, Andy; Weiner, Joel H.; Li, Liang

    2013-01-01

    Graphical abstract: -- Highlights: •Investigating a strategy of reciprocal isotope labeling of comparative samples. •Filtering out incorrect peptide identification or quantification values. •Analyzing the proteome changes of E. coli cells grown aerobically or anaerobically. •Presenting guidelines for reciprocal labeling experimental design. -- Abstract: Due to limited sample amounts, instrument time considerations, and reagent costs, only a small number of replicate experiments are typically performed for quantitative proteome analyses. Generation of reproducible data that can be readily assessed for consistency within a small number of datasets is critical for accurate quantification. We report our investigation of a strategy using reciprocal isotope labeling of two comparative samples as a tool for determining proteome changes. Reciprocal labeling was evaluated to determine the internal consistency of quantified proteome changes from Escherichia coli grown under aerobic and anaerobic conditions. Qualitatively, the peptide overlap between replicate analyses of the same sample and reverse labeled samples were found to be within 8%. Quantitatively, reciprocal analyses showed only a slight increase in average overall inconsistency when compared with replicate analyses (1.29 vs. 1.24-fold difference). Most importantly, reverse labeling was successfully used to identify spurious values resulting from incorrect peptide identifications and poor peak fitting. After removal of 5% of the peptide data with low reproducibility, a total of 275 differentially expressed proteins (>1.50-fold difference) were consistently identified and were then subjected to bioinformatics analysis. General considerations and guidelines for reciprocal labeling experimental design and biological significance of obtained results are discussed

  18. Adsorption methods for hydrogen isotope storage on zeolite sieves

    International Nuclear Information System (INIS)

    Cristescu, Ioana; Cristescu, Ion; Vasut, Felicia; Brad, Sebastian; Lazar, Alin

    2001-01-01

    Adsorption molecular sieves and activated carbon were used for hydrogen isotopes. The adsorption process proceeds at liquid nitrogen and liquid hydrogen temperatures. The synthetic zeolites have similar properties as natural zeolites, but they have a regular pore structure and affinity for molecules of different size with defined shapes. Experimental results obtained at liquid nitrogen and liquid hydrogen temperatures evidenced the efficient behavior of the activated carbon and zeolite sieves for hydrogen isotope temporary storage. (authors)

  19. The application of isotopic dating methods for prospection and exploration of nuclear raw material

    International Nuclear Information System (INIS)

    Komlev, L.V.; Anderson, E.B.

    1977-01-01

    Among the geological and geochemical methods for prospecting and searching the nuclear raw material, the isotope-dating methods determine the most important search criterion - the time of the ore-forming. The elaboration and use of these methods in uranium-ore regions reveal a series of geochemical epochs of uranium and thorium accumulation connected naturally with the history of geological evolution of the earth crust. The isotope-dating methods enable with confidence to establish the stages of tectono-magmatic activity resulting in the redistribution and the local concentration of uranium. The wide use of isotopic methods is a necessary condition for reasonable trends of the modern geological exploration [ru

  20. Extracorporeal adsorption therapy: A Method to improve targeted radiation delivered by radiometal-labeled monoclonal antibodies

    International Nuclear Information System (INIS)

    Nemecek, Eneida R.; Green, Damian J.; Fisher, Darrell R.; Pagal, John M.; Lin, Yukang; Gopal, A. K.; Durack, Lawrence D.; Rajendran, Joseph G.; Wilbur, D. S.; Nilsson, Rune; Sandberg, Bengt; Press, Oliver W.

    2008-01-01

    Many investigators have demonstrated the ability to treat hematologic malignancies with radiolabeled monoclonal antibodies targeting hematopoietic antigens such as anti-CD20 and anti-CD45. [1-5] Although the remission rates achieved with radioimmunotherapy (RIT) are relatively high, many patients subsequently relapse presumably due to suboptimal delivery of enough radiation to eradicate the malignancy. The dose-response of leukemia and lymphoma to radiation has been proven. Substantial amounts of radiation can be delivered by RIT if followed by hematopoietic cell transplantation to rescue the bone marrow from myeloablation.[ref] However, the maximum dose of RIT that can be used is still limited by toxicity to normal tissues affected by nonspecific delivery of radiation. Efforts to improve RIT focus on improving the therapeutic ratios of radiation in target versus non-target tissues by removing the fraction of radioisotope that fails to bind to target tissues and circulates freely in the bloodstream perfusing non-target tissues. Our group and others have explored several alternatives for removal of unbound circulating antibody. [refs] One such method, extracorporeal adsorption therapy (ECAT) consists of removing unbound antibody by a method similar to plasmapheresis after critical circulation time and distribution of antibody into target tissues have been achieved. Preclinical studies of ECAT in murine xenograft models demonstrated significant improvement in therapeutic ratios of radioactivity. Chen and colleagues demonstrated that a 2-hour ECAT procedure could remove 40 to 70% of the radioactivity from liver, lung and spleen. [ref] Although isotope concentration in the tumor was initially unaffected, a 50% decrease was noted approximately 36 hours after the procedure. This approach was also evaluated in a limited phase I pilot study of patients with refractory B-cell lymphoma. [ref] After radiographic confirmation of tumor localization of a test dose of anti-CD20

  1. Methods for labeling .beta.-amyloid plaques and neurofibrillary tangles

    Science.gov (United States)

    Barrio, Jorge R.; Petric, Andrej; Satyamurthy, Nagichettiar; Small, Gary W.; Cole, Gregory M.; Huang, Sung-Cheng

    2001-01-01

    A method for labeling .beta.-amyloid plaques and neurofibrillary tangles in vivo and in vitro, comprises contacting a compound of formula (I): ##STR1## with mammalian tissue. In formula (I), R.sub.1 is selected from the group consisting of --C(O)-alkyl, --C(O)-alkylenyl-R.sub.4, --C(O)O-alkyl, --C(O)O-alkylenyl-R.sub.4, --C.dbd.C(CN).sub.2 -alkyl, --C.dbd.C(CN).sub.2 -alkylenyl-R.sub.4 , ##STR2## R.sub.4 is a radical selected from the group consisting of alkyl, substituted alkyl, aryl and substituted aryl; R.sub.5, is a radical selected from the group consisting of --NH.sub.2, --OH, --SH, --NH-alkyl, --NHR.sub.4, --NH-alkylenyl-R.sub.4, --O-alkyl, --O-alkylenyl-R.sub.4, --S-alkyl, and --S-alkylenyl-R.sub.4 ; R.sub.6 is a radical selected from the group consisting of --CN, --COOH, --C(O)O-alkyl, --C(O)O-alkylenyl-R.sub.4, --C(O)-alkyl, --C(O)-alkylenyl-R.sub.4, --C(O)-halogen, --C(O)NH , --C(O)NH-alkyl, --C(O)NH-alkylenyl-R.sub.4 ; R.sub.7 is a radical selected from the group consisting of O, NH, and S; and R.sub.8 is N, O or S. R.sub.2 and R.sub.3 are each independently selected from the group consisting of alkyl and alkylenyl-R.sub.10, wherein R.sub.10 is selected from the group consisting of --OH, --OTs, halogen, spiperone, spiperone ketal and spiperone-3-yl. Alternatively, R.sub.2 and R.sub.3 together form a heterocyclic ring, optionally substituted with at least one radical selected from the group consisting of alkyl, alkoxy, OH, OTs, halogen, alkylenyl-R.sub.10, carbonyl, spiperone, spiperone ketal and spiperone-3-yl. In the compounds of formula (I), one or more of the hydrogen, halogen or carbon atoms can, optionally, be replaced with a radiolabel.

  2. Hydrogen isotope recovering and reutilizing method and its device

    International Nuclear Information System (INIS)

    Ide, Takahiro.

    1988-01-01

    Purpose: To enable safety and convenient recovery and reutilization of gaseous tritium and other hydrogen isotopes. Constitution: Two kinds of metal hydrides different from each other in the dissociation pressure at an identical temperature are combined, in which a metal hydride of higher dissociation pressure is used for recovery and reutilization for most portion of gaseous hydrogen isotope gases, while the metal hydride of lower dissociation pressure is used for the recovery and reutilization of the remaining gaseous hydrogen isotopes. This enables to extremely lower the concentration of the remaining gaseous hydrogen isotopes, that is, the concentration of tritium in the recoverying system. In addition, since the heating temperature required for releasing the gaseous hydrogen isotopes absorbed in both of the metal hydrides is within such a range as causing no problem for the permeation of the gaseous hydrogen isotopes, there is no requirement for the countermeasure to tritium permeation or the facility for recovering permeated tritium and there is no problem for the material degradation due to the heating at high temperature. (Kawakami, Y.)

  3. Semiempirical method to determine the uranium isotopic compositions

    International Nuclear Information System (INIS)

    Tegas Sutondo

    2008-01-01

    In a nuclear reactor design calculation, some variations of U 235 enrichment are commonly needed. This will affect the isotopic compositions of the 3 main uranium isotopes i.e. U 234 , U 235 and U 238 for the respective enrichment. Due to the limited compositions data available, it is urgent to make an approximate way that can be used to determine the compositions of the 3 isotopes, for the desired enrichments. This paper presents the theoretical background used for constructing a semi empirical formula to estimate the composition of the 3 uranium isotopes as a function of U 235 enrichment, obtained based on the measurement data available. Based on the available data, and the lack of compositions data within the enrichment range between 3.5 % and around 12 %, it is concluded that 2 separate linear equations i.e. for ≤ 3.5 % and ≥ 3.5 % might be needed for U 235 isotope. For the U 234 isotope, a polynomial equation of 4 th order is well suited to be used for the whole range of enrichment between 0.711 % and 20 %, whilst for higher enrichment (> 20 %), a power function seems to give a better approach. The composition of U 238 can then be determined from the U 235 and U 234 composition at the desired enrichment of U 235 . (author)

  4. Modification degrees at specific sites on heparan sulphate: an approach to measure chemical modifications on biological molecules with stable isotope labelling

    Science.gov (United States)

    Wu, Zhengliang L.; Lech, Miroslaw

    2005-01-01

    Chemical modification of biological molecules is a general mechanism for cellular regulation. A quantitative approach has been developed to measure the extent of modification on HS (heparan sulphates). Sulphation on HS by sulphotransferases leads to variable sulphation levels, which allows cells to tune their affinities to various extracellular proteins, including growth factors. With stable isotope labelling and HPLC-coupled MS, modification degrees at various O-sulphation sites could be determined. A bovine kidney HS sample was first saturated in vitro with 34S by an OST (O-sulphotransferase), then digested with nitrous acid and analysed with HPLC-coupled MS. The 34S-labelled oligosaccharides were identified based on their unique isotope clusters. The modification degrees at the sulphotransferase recognition sites were obtained by calculating the intensities of isotopic peaks in the isotope clusters. The modification degrees at 3-OST-1 and 6-OST-1 sites were examined in detail. This approach can also be used to study other types of chemical modifications on biological molecules. PMID:15743272

  5. A revised method of labeling mouse IgG with yttrium-90

    International Nuclear Information System (INIS)

    Arbab, A.S.; Koizumi, Kiyoshi; Araki, Tsutomu

    1996-01-01

    We report the successful labeling of mouse IgG with yttrium-90 (Y-90) using isothiocyanatobenzyl-ethylenediaminetetraacetic acid (SCN-Bz-EDTA) as a chelating agent and compared the result with labeling by indium-111 (In-111). After conjugating IgG with SCN-Bz-EDTA, a predetermined volume of conjugated IgG was mixed with different volumes of either Y-90 or In-111 acetate and incubated at 37degC. Labeling efficiency was assessed at specific intervals upto 3 hr. After 3 hr, the mixtures were challenged with Na 2 EDTA to evaluate the transchelation of labeled Y-90 or In-111. All mixtures showed labeling efficiency of around 50% with Y-90 and the leveling was fairly preserved even after Na 2 EDTA challenge. However, labeling with In-111 was unsuccessful when conjugated IgG was not separated from the unconjugated form. When separated, however, In-111 showed more than 80% labeling efficiency though labeling with In-111 could not tolerate Na 2 EDTA challenge. In conclusion IgG was efficiently labeled by Y-90 using SCN-Bz-EDTA though labeling with In-111 showed some problems associated with this method. (author)

  6. CO2 production in animals: analysis of potential errors in the doubly labeled water method

    International Nuclear Information System (INIS)

    Nagy, K.A.

    1979-03-01

    Laboratory validation studies indicate that doubly labeled water ( 3 HH 18 O and 2 HH 18 O) measurements of CO 2 production are accurate to within +-9% in nine species of mammals and reptiles, a bird, and an insect. However, in field studies, errors can be much larger under certain circumstances. Isotopic fraction of labeled water can cause large errors in animals whose evaporative water loss comprises a major proportion of total water efflux. Input of CO 2 across lungs and skin caused errors exceeding +80% in kangaroo rats exposed to air containing 3.4% unlabeled CO 2 . Analytical errors of +-1% in isotope concentrations can cause calculated rates of CO 2 production to contain errors exceeding +-70% in some circumstances. These occur: 1) when little decline in isotope concentractions has occured during the measurement period; 2) when final isotope concentrations closely approach background levels; and 3) when the rate of water flux in an animal is high relative to its rate of CO 2 production. The following sources of error are probably negligible in most situations: 1) use of an inappropriate equation for calculating CO 2 production, 2) variations in rates of water or CO 2 flux through time, 3) use of H 2 O-18 dilution space as a measure of body water volume, 4) exchange of 0-18 between water and nonaqueous compounds in animals (including excrement), 5) incomplete mixing of isotopes in the animal, and 6) input of unlabeled water via lungs and skin. Errors in field measurements of CO 2 production can be reduced to acceptable levels (< 10%) by appropriate selection of study subjects and recapture intervals

  7. Pb and Sr isotopic compositions of ancient pottery: a method to discriminate production sites

    International Nuclear Information System (INIS)

    Zhang Xun; Chen Jiangfeng; Ma Lin; He Jianfeng; Wang Changsui; Qiu Ping

    2004-01-01

    The discriminating of production sites of ancient pottery samples using multi-isotopic systematics was described. Previous work has proven that Pb isotopic ratios can be used for discriminating the production sites of ancient pottery under certain conditions. The present work suggests that although Nd isotopic ratios are not sensitive to the production sites of ancient pottery, Sr isotopic ratios are important for the purpose. Pb isotopic ratios are indistinguishable for the pottery excavated from the Jiahu relict, Wuyang, Henan Province and for famous Qin Terra-cotta Figures. But, the 87 Sr/ 86 Sr ratios for the former (about 0.715) are significantly lower than that of the latter (0.717-0.718). The authors concluded that a combined use of Pb and Sr isotopes would be a more powerful method for discriminating the production site of ancient pottery. (authors)

  8. A simple method for stem cell labeling with fluorine 18

    International Nuclear Information System (INIS)

    Ma Bing; Hankenson, Kurt D.; Dennis, James E.; Caplan, Arnold I.; Goldstein, Steven A.; Kilbourn, Michael R.

    2005-01-01

    Hexadecyl-4-[ 18 F]fluorobenzoate ([ 18 F]HFB), a long chain fluorinated benzoic acid ester, was prepared in a one-step synthesis by aromatic nucleophilic substitution of [ 18 F]fluoride ion on hexadecyl-4-(N,N,N-trimethylammonio)benzoate. The radiolabeled ester was obtained in good yields (52% decay corrected) and high purity (97%). [ 18 F]HFB was used to radiolabel rat mesenchymal stem cells (MSCs) by absorption into cell membranes. MicroPET imaging of [ 18 F]HFB-labeled MSCs following intravenous injection into the rat showed the expected high and persistent accumulation of radioactivity in the lungs. [ 18 F]HFB is thus simple to prepare and uses labeling agent for short-term distribution studies of injected stem cells

  9. A simple method for stem cell labeling with fluorine 18

    Energy Technology Data Exchange (ETDEWEB)

    Ma Bing [Department of Radiology, Division of Nuclear Medicine, University of Michigan Medical School, Ann Arbor, MI 48109 (United States); Hankenson, Kurt D. [Department of Biology, Case Western Reserve University, Cleveland, OH 44106 (United States); Dennis, James E. [Department of Biology, Case Western Reserve University, Cleveland, OH 44106 (United States); Caplan, Arnold I. [Department of Biology, Case Western Reserve University, Cleveland, OH 44106 (United States); Goldstein, Steven A. [Department of Orthopaedic Surgery, University of Michigan Medical School, Ann Arbor, MI 48109 (United States); Kilbourn, Michael R. [Department of Radiology, Division of Nuclear Medicine, University of Michigan Medical School, Ann Arbor, MI 48109 (United States)

    2005-10-01

    Hexadecyl-4-[{sup 18}F]fluorobenzoate ([{sup 18}F]HFB), a long chain fluorinated benzoic acid ester, was prepared in a one-step synthesis by aromatic nucleophilic substitution of [{sup 18}F]fluoride ion on hexadecyl-4-(N,N,N-trimethylammonio)benzoate. The radiolabeled ester was obtained in good yields (52% decay corrected) and high purity (97%). [{sup 18}F]HFB was used to radiolabel rat mesenchymal stem cells (MSCs) by absorption into cell membranes. MicroPET imaging of [{sup 18}F]HFB-labeled MSCs following intravenous injection into the rat showed the expected high and persistent accumulation of radioactivity in the lungs. [{sup 18}F]HFB is thus simple to prepare and uses labeling agent for short-term distribution studies of injected stem cells.

  10. Phosphorus availability to ryegrass from urban sewage sludges assessed by isotopic labeling and dilution technique, effect of irradiation

    International Nuclear Information System (INIS)

    El-Motaium, R.A.; Morel, C.

    2007-01-01

    Urban sewage sludge are widely used as alternative P input because of their high P content. Irradiation of sewage sludge is developed in Egypt to make safe spreading of sludge on agricultural fields. Five sludges were sampled from representative urban wastewater treatment plants located in Cairo, Egypt. Sub samples were irradiated using gamma radiation at 6 KGy dose. A pot experiment was conducted under plastic greenhouse using sandy P-deficient soil. The soil was homogeneously labelled with a radioactive solution of carrier-free 32 PO 4 (about 3 MBq kg / soil) to determine the contribution of the different sources of P in plant nutrition. Sludges were applied at 50 mg P/ kg soil. In addition, a control treatment was also carried out to analyze sludge P availability in comparison to a water soluble mineral fertilizer, i.e. 50 mg P/kg soil was applied as commercial monocalcium phosphate (SSP). A reference treatment was included without any P application but the radioactive solution was added. One gram of ryegrass seeds (Lolium multiflorium) was sown in each pot. The aerial parts of the ryegrass were harvested four times, every 3 weeks. Analysis of all harvests included 32 P and total P content was used to determine the respective contribution of soil and sludge to plant P nutrition and the plant available P (L value) using the isotopic dilution principle. The ratio of radioactive P to non-radioactive P, i.e. isotopic composition, was calculated. The total P content in the different urban sewage sludge was ranged from 6.2 to 13.8 g P/ kg and affected by irradiation. Because the soil was extremely P deficient (L value=0.3 mg P/ kg soil), P derived from seeds represented 99% of the P taken up when no P was applied and 87% when applying 50 mg P-SSP/ kg. After correcting the contribution of ryegrass seeds, sludge P contributed by 98% to plant nutrition and 7.2% of the applied P was recovered. The sludge type and irradiation factors did not significantly affect sludge P

  11. Design study of fuel circulating system using Pd-alloy membrane isotope separation method

    International Nuclear Information System (INIS)

    Naito, T.; Yamada, T.; Yamanaka, T.; Aizawa, T.; Kasahara, T.; Nishikawa, M.; Asami, N.

    1980-01-01

    Design study on the fuel circulating system (FCS) for a tokamak experimental fusion reactor (JXFR) has been carried out to establish the system concept, to plan the development program, and to evaluate the feasibility of diffusion system. The FCS consists of main vacuum system, fuel gas refiners, isotope separators, fuel feeders, and auxiliary systems. In the system design, Pd-alloy membrane permeation method is adopted for fuel refining and isotope separating. All impurities are effectively removed and hydrogen isotopes are sufficiently separated by Pd-alloy membrane. The isotope separation system consists of 1st (47 separators) and 2nd (46 separators) cascades for removing protium and separating deuterium, respectively. In the FCS, while cryogenic distillation method appears to be practicable, Pd-alloy membrane diffusion method is attractive for isotope separation and refining of fuel gas. The choice will have to be based on reliability, economic, and safety analyses

  12. Kinetic investigation of heterogeneous catalytic reactions by means of the kinetic isotope method

    Energy Technology Data Exchange (ETDEWEB)

    Bauer, F; Dermietzel, J [Akademie der Wissenschaften der DDR, Leipzig. Zentralinstitut fuer Isotopen- und Strahlenforschung

    1978-09-01

    The application of the kinetic isotope method to heterogeneous catalytic processes is possible for surface compounds by using the steady-state relation. However, the characterization of intermediate products becomes ambiguous if sorption rates are of the same order of magnitude as surface reactions rates. The isotopic exchange reaction renders possible the estimation of sorption rates.

  13. SORIOS – A method for evaluating and selecting environmental certificates and labels

    DEFF Research Database (Denmark)

    Kikkenborg Pedersen, Dennis; Dukovska-Popovska, Iskra; Ola Strandhagen, Jan

    2012-01-01

    This paper presents a general method for evaluating and selecting environmental certificates and labels for companies to use on products and services. The method is developed based on a case study using a Grounded Theory approach. The result is a generalized six-step method that features an initial...... searching strategy and an evaluation model that weighs the prerequisites, rewards and the organization of certificate or label against the strategic needs of a company....

  14. Stable isotope methods: The effect of gut contents on isotopic ratios of zooplankton

    Science.gov (United States)

    Hill, J. M.; McQuaid, C. D.

    2011-05-01

    In the past decade there has been an increased awareness of the potential for methodological bias resulting from multiple pre-analytical procedures in foodweb interpretations based on stable isotope techniques. In the case of small organisms, this includes the effect of gut contents on whole body signatures. Although gut contents may not reflect actual assimilation, their carbon and nitrogen values will be isotopically lighter than after the same material has been assimilated. The potential skewing of isotopic ratios in whole organism samples is especially important for aquatic environments as many studies involve trophic relationships among small zooplankton. This is particularly important in pelagic waters, where herbivorous zooplankton comprise small taxa. Hence this study investigated the effect of gut contents on the δ13C and δ15N ratios of three size classes of zooplankton (1.0-2.0, 2.0-4.0 and >4.0 mm) collected using bongo net tows in the tropical waters of the south-west Indian Ocean. Animals were collected at night, when they were likely to be feeding, sieved into size classes and separated into genera. We focused on Euphausia spp which dominated zooplankton biomass. Three treatment types were processed: bulk animals, bulk animals without guts and tail muscle from each size class at 10 bongo stations. The δ15N ratios were influenced by zooplankton size class, presumably reflecting ontogenetic changes in diet. ANOVA post hoc results and correlations in δ15N signatures among treatments suggest that gut contents may not affect overall nitrogen signatures of Euphausia spp., but that δ13C signatures may be significantly altered by their presence. Carbon interpretations however, were complicated by potential effects of variation in chitin, lipids and metabolism among tissues and the possibility of opportunistic omnivory. Consequently we advocate gut evacuation before sacrifice in euphausiids if specific tissue dissection is impractical and recommend

  15. Preliminary results of oxygen isotope ratio measurement with a particle-gamma coincidence method

    Energy Technology Data Exchange (ETDEWEB)

    Borysiuk, Maciek, E-mail: maciek.borysiuk@pixe.lth.se; Kristiansson, Per; Ros, Linus; Abdel, Nassem S.; Elfman, Mikael; Nilsson, Charlotta; Pallon, Jan

    2015-04-01

    The possibility to study variations in the oxygen isotopic ratio with photon tagged nuclear reaction analysis (pNRA) is evaluated in the current work. The experiment described in the article was performed at Lund Ion Beam Analysis Facility (LIBAF) with a 2 MeV deuteron beam. Isotopic fractionation of light elements such as carbon, oxygen and nitrogen is the basis of many analytical tools in hydrology, geology, paleobiology and paleogeology. IBA methods provide one possible tool for measurement of isotopic content. During this experimental run we focused on measurement of the oxygen isotopic ratio. The measurement of stable isotopes of oxygen has a number of applications; the particular one driving the current investigation belongs to the field of astrogeology and specifically evaluation of fossil extraterrestrial material. There are three stable isotopes of oxygen: {sup 16}O, {sup 17}O and {sup 18}O. We procured samples highly enriched with all three isotopes. Isotopes {sup 16}O and {sup 18}O were easily detected in the enriched samples, but no significant signal from {sup 17}O was detected in the same samples. The measured yield was too low to detect {sup 18}O in a sample with natural abundances of oxygen isotopes, at least in the current experimental setup, but the spectral line from the reaction with {sup 16}O was clearly visible.

  16. Method for separating the isotopes of a chemical element

    International Nuclear Information System (INIS)

    Devienne, F.M.

    1977-01-01

    A beam of positive or negative primary ions of at least one compound of a chemical element is accelerated in order to pass through collision boxes placed in series. As a result of inelastic collisions of the ions with the molecules of a neutral target gas within each collision box, a given percentage of primary ions is dissociated into at least two fragments, one of which is a secondary ion in the form of at least two isotopic species. The collision boxes are brought to a potential V 2 so as to trap preferentially one isotopic species which is condensed within each box. 15 claims, 4 figures

  17. Methyl-coenzyme M reductase from methanogenic archaea: isotope effects on label exchange and ethane formation with the homologous substrate ethyl-coenzyme M.

    Science.gov (United States)

    Scheller, Silvan; Goenrich, Meike; Thauer, Rudolf K; Jaun, Bernhard

    2013-10-09

    Ethyl-coenzyme M (CH3CH2-S-CH2CH2-SO3(-), Et-S-CoM) serves as a homologous substrate for the enzyme methyl-coenzyme M reductase (MCR) resulting in the product ethane instead of methane. The catalytic reaction proceeds via an intermediate that already contains all six C-H bonds of the product. Because product release occurs after a second, rate-limiting step, many cycles of intermediate formation and reconversion to substrate occur before a substantial amount of ethane is released. In deuterated buffer, the intermediate becomes labeled, and C-H activation in the back reaction rapidly leads to labeled Et-S-CoM, which enables intermediate formation to be detected. Here, we present a comprehensive analysis of this pre-equilibrium. (2)H- and (13)C-labeled isotopologues of Et-S-CoM were used as the substrates, and the time course of each isotopologue was followed by NMR spectroscopy. A kinetic simulation including kinetic isotope effects allowed determination of the primary and α- and β-secondary isotope effects for intermediate formation and for the C-H/C-D bond activation in the ethane-containing intermediate. The values obtained are in accordance with those found for the native substrate Me-S-CoM (see preceding publication, Scheller, S.; Goenrich, M.; Thauer, R. K.; Jaun, B. J. Am. Chem. Soc. 2013, 135, DOI: 10.1021/ja406485z) and thus imply the same catalytic mechanism for both substrates. The experiment by Floss and co-workers, demonstrating a net inversion of configuration to chiral ethane with CH3CDT-S-CoM as the substrate, is compatible with the observed rapid isotope exchange if the isotope effects measured here are taken into account.

  18. A Graphite Isotope Ratio Method: A Primer on Estimating Plutonium Production in Graphite Moderated Reactors

    International Nuclear Information System (INIS)

    Gesh, Christopher J.

    2004-01-01

    The Graphite Isotope Ratio Method (GIRM) is a technique used to estimate the total plutonium production in a graphite-moderated reactor. The cumulative plutonium production in that reactor can be accurately determined by measuring neutron irradiation induced isotopic ratio changes in certain impurity elements within the graphite moderator. The method does not require detailed knowledge of a reactor's operating history, although that knowledge can decrease the uncertainty of the production estimate. The basic premise of the Graphite Isotope Ratio Method is that the fluence in non-fuel core components is directly related to the cumulative plutonium production in the nuclear fuel

  19. Magnetic resonance studies of isotopically labeled paramagnetic proteins: (2FE-2S) ferredoxins

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, H.; Xia, B.; Chae, Y.K.; Westler, W.M.; Markley, J.L. [Univ. of Wisconsin, Madison, WI (United States)

    1994-12-01

    Recent developments in NMR spectroscopy, especially multidimensional, multinuclear NMR techniques, have made NMR the most versatile tool available for studying protein structure and function in solution. Unlike diamagnetic proteins, paramagnetic proteins contain centers with unpaired electrons. These unpaired electrons interact with magnetic nuclei either through chemical bonds by a contact mechanism or through space by a pseudocontact mechanism. Such interactions make the acquisition and analysis of NMR spectra of paramagnetic proteins more challenging than those of diamagnetic proteins. Some NMR signals from paramagnetic proteins are shifted outside the chemical shift region characteristic of diamagnetic proteins; these {open_quotes}hyperfine-shifted{close_quotes} resonances originate from nuclei that interact with unpaired electrons from the paramagnetic center. The large chemical shift dispersion in spectra of paramagnetic proteins makes it difficult to excite the entire spectral window and leads to distortions in the baseline. Interactions with paramagnetic centers shorten T{sub 1} and T{sub 2} relaxation times of nuclei; the consequences are line broadening and lower spectral sensitivity. Scalar (through bond) and dipolar (through space) interactions between pairs of nuclei are what give rise to crosspeak signals in multi-dimensional NMR spectra of small diamagnetic proteins. When such interactions involve a nucleus that is strongly relaxed by interaction with a paramagnetic center, specialized methods may be needed for its detection or it may be completely undetectable by present nD NMR methods.

  20. COMBINING SOURCES IN STABLE ISOTOPE MIXING MODELS: ALTERNATIVE METHODS

    Science.gov (United States)

    Stable isotope mixing models are often used to quantify source contributions to a mixture. Examples include pollution source identification; trophic web studies; analysis of water sources for soils, plants, or water bodies; and many others. A common problem is having too many s...

  1. Solution of the isotopic depletion equation using decomposition method and analytical solution

    Energy Technology Data Exchange (ETDEWEB)

    Prata, Fabiano S.; Silva, Fernando C.; Martinez, Aquilino S., E-mail: fprata@con.ufrj.br, E-mail: fernando@con.ufrj.br, E-mail: aquilino@lmp.ufrj.br [Coordenacao dos Programas de Pos-Graduacao de Engenharia (PEN/COPPE/UFRJ), RJ (Brazil). Programa de Engenharia Nuclear

    2011-07-01

    In this paper an analytical calculation of the isotopic depletion equations is proposed, featuring a chain of major isotopes found in a typical PWR reactor. Part of this chain allows feedback reactions of (n,2n) type. The method is based on decoupling the equations describing feedback from the rest of the chain by using the decomposition method, with analytical solutions for the other isotopes present in the chain. The method was implemented in a PWR reactor simulation code, that makes use of the nodal expansion method (NEM) to solve the neutron diffusion equation, describing the spatial distribution of neutron flux inside the reactor core. Because isotopic depletion calculation module is the most computationally intensive process within simulation systems of nuclear reactor core, it is justified to look for a method that is both efficient and fast, with the objective of evaluating a larger number of core configurations in a short amount of time. (author)

  2. Solution of the isotopic depletion equation using decomposition method and analytical solution

    International Nuclear Information System (INIS)

    Prata, Fabiano S.; Silva, Fernando C.; Martinez, Aquilino S.

    2011-01-01

    In this paper an analytical calculation of the isotopic depletion equations is proposed, featuring a chain of major isotopes found in a typical PWR reactor. Part of this chain allows feedback reactions of (n,2n) type. The method is based on decoupling the equations describing feedback from the rest of the chain by using the decomposition method, with analytical solutions for the other isotopes present in the chain. The method was implemented in a PWR reactor simulation code, that makes use of the nodal expansion method (NEM) to solve the neutron diffusion equation, describing the spatial distribution of neutron flux inside the reactor core. Because isotopic depletion calculation module is the most computationally intensive process within simulation systems of nuclear reactor core, it is justified to look for a method that is both efficient and fast, with the objective of evaluating a larger number of core configurations in a short amount of time. (author)

  3. Two-point vs multipoint sample collection for the analysis of energy expenditure by use of the doubly labeled water method

    International Nuclear Information System (INIS)

    Welle, S.

    1990-01-01

    Energy expenditure over a 2-wk period was determined by the doubly labeled water (2H2(18)O) method in nine adults. When daily samples were analyzed, energy expenditure was 2859 +/- 453 kcal/d (means +/- SD); when only the first and last time points were considered, the mean calculated energy expenditure was not significantly different (2947 +/- 430 kcal/d). An analysis of theoretical cases in which isotope flux is not constant indicates that the multipoint method can cause errors in the calculation of average isotope fluxes, but these are generally small. Simulations of the effect of analytical error indicate that increasing the number of replicates on two points reduces the impact of technical errors more effectively than does performing single analyses on multiple samples. It appears that generally there is no advantage to collecting frequent samples when the 2H2(18)O method is used to estimate energy expenditure in adult humans

  4. Labelled compounds. (Pt. B)

    International Nuclear Information System (INIS)

    Buncel, E.; Jones, J.R.

    1991-01-01

    Since the end of World War II there has been a tremendous increase in the number of compounds that have been synthesized with radioactive or stable isotopes. They have found application in many diverse fields, so much so, that hardly a single area in pure and applied science has not benefited. Not surprisingly it has been reflected in appearance of related publications. The early proceedings of the Symposia on Advances in Trace Methodology were soon followed by various Euratom sponsored meetings in which methods of preparing and storing labelled compounds featured prominently. In due course a resurgence of interest in stable isotopes, brought about by their greater availability (also lower cost) and partly by development of new techniques such as gas chromatography - mass spectrometry (gc-ms), led to the publication of proceedings of several successful conferences. More recently conferences dealing with the synthesis and applications of isotopes and isotopically labelled compounds have been established on a regular basis. In addition to the proceedings of conferences and journal publications individuals left their mark by producing definitive texts, usually on specific nuclides. Only the classic two volume publication of Murray and Williams (Organic syntheses with isotopes, New York 1985), now over 30 years old and out of print, attempted to do justice to several nuclides. With the large amount of work that has been undertaken since then it seems unlikely that an updated edition could be produced. The alternative strategy was to ask scientists currently active to review specific areas and this is the approach adopted in the present series of monographs. In this way it is intended to cover the broad advances that have been made in the synthesis and applications of isotopes and isotopically labelled compounds in the physical and biomedical sciences. (author). refs.; figs.; tabs

  5. Exploring the nitrogen ingestion of aphids--a new method using electrical penetration graph and (15N labelling.

    Directory of Open Access Journals (Sweden)

    Franziska Kuhlmann

    Full Text Available Studying plant-aphid interactions is challenging as aphid feeding is a complex process hidden in the plant tissue. Here we propose a combination of two well established methods to study nutrient acquisition by aphids focusing on the uptake of isotopically labelled nitrogen ((15N. We combined the Electrical Penetration Graph (EPG technique that allows detailed recording of aphid feeding behaviour and stable isotope ratio mass spectrometry (IRMS to precisely measure the uptake of nitrogen. Bird cherry-oat aphids Rhopalosiphum padi L. (Hemiptera, Aphididae fed for 24 h on barley plants (Hordeum vulgare L., cultivar Lina, Poaceae that were cultivated with a (15N enriched nutrient solution. The time aphids fed in the phloem was strongly positive correlated with their (15N uptake. All other single behavioural phases were not correlated with (15N enrichment in the aphids, which corroborates their classification as non-feeding EPG phases. In addition, phloem-feeding and (15N enrichment of aphids was divided into two groups. One group spent only short time in the phloem phase and was unsuccessful in nitrogen acquisition, while the other group displayed longer phloem-feeding phases and was successful in nitrogen acquisition. This suggests that several factors such as the right feeding site, time span of feeding and individual conditions play a role for the aphids to acquire nutrients successfully. The power of this combination of methods for studying plant-aphid interactions is discussed.

  6. Technetium 99-m labeled radio-diagnostic agents employing stannous tartrate and method of preparation

    International Nuclear Information System (INIS)

    Molinski, V.J.; Wilczewski, J.A.

    1976-01-01

    A method of preparing improved technetium-99m labeled radiodiagnostic agents by reducing technetium-99m with stannous tartrate. Such radiodiagnostic agents are useful in scintigraphic examinations of the bone and lung. 31 claims, no drawings

  7. A comparative study on the iodine-labeled methods of protein and polypeptide

    International Nuclear Information System (INIS)

    Li Huaifen; Niu Huisheng; Yuan Mingyue; Yu Jinghua

    1994-01-01

    There are three methods: chloramine-T, Iodogen and lactoperoxidase(LPO). 125 I-ACTH, 125 I-insulin and 125 I-HSA are prepared by these techniques. The results show that lactoperoxidase is isolated and purified from fresh milk, meanwhile, the enzyme is used in experiments of 125 I-labeled protein, peptide hormone and mono-clone antibody, etc. LPO is a very successful method for it's mild, complete reaction, controllable, high labelling yield, higher purity of iodine-labeled compound and so on. It remains biological activation and stable character more than other two techniques

  8. A comparative study on the iodine-labeled methods of protein and polypeptide

    Energy Technology Data Exchange (ETDEWEB)

    Huaifen, Li; Huisheng, Niu; Mingyue, Yuan; Jinghua, Yu [Chinese Academy of Medical Sciences, Tianjin (China). Inst. of Radiation Medicine

    1994-02-01

    There are three methods: chloramine-T, Iodogen and lactoperoxidase(LPO). [sup 125]I-ACTH, [sup 125]I-insulin and [sup 125]I-HSA are prepared by these techniques. The results show that lactoperoxidase is isolated and purified from fresh milk, meanwhile, the enzyme is used in experiments of [sup 125]I-labeled protein, peptide hormone and mono-clone antibody, etc. LPO is a very successful method for it's mild, complete reaction, controllable, high labelling yield, higher purity of iodine-labeled compound and so on. It remains biological activation and stable character more than other two techniques.

  9. Isotopic measurements (C,N,O) of detonation soot produced from labeled and unlabeled Composition B-3 indicate source of solid carbon residues

    Science.gov (United States)

    Podlesak, David; Manner, Virginia; Amato, Ronald; Dattelbaum, Dana; Gusavsen, Richard; Huber, Rachel

    2017-06-01

    Detonation of HE is an exothermic process whereby metastable complex molecules are converted to simple stable molecules such as H2 O, N2, CO, CO2, and solid carbon. The solid carbon contains various allotropes such as detonation nanodiamonds, graphite, and amorphous carbon. It is well known that certain HE formulations such as Composition B (60% RDX, 40% TNT) produce greater amounts of solid carbon than other more oxygen-balanced formulations. To develop a greater understanding of how formulation and environment influence solid carbon formation, we synthesized TNT and RDX with 13 C and 15 N at levels slightly above natural abundance levels. Synthesized RDX and TNT were mixed at a ratio of 60:40 to form Composition B and solid carbon residues were collected from detonations of isotopically-labeled as well as un-labelled Composition B. The raw HE and detonation residues were analyzed isotopically for C, N, O isotopic compositions. We will discuss differences between treatments groups as a function of formulation and environment. LA-UR - 17-21266.

  10. Isotope dilution liquid chromatography - mass spectrometry methods for fat- and water-soluble vitamins in nutritional formulations.

    Science.gov (United States)

    Phinney, Karen W; Rimmer, Catherine A; Thomas, Jeanice Brown; Sander, Lane C; Sharpless, Katherine E; Wise, Stephen A

    2011-01-01

    Vitamins are essential to human health, and dietary supplements containing vitamins are widely used by individuals hoping to ensure they have adequate intake of these important nutrients. Measurement of vitamins in nutritional formulations is necessary to monitor regulatory compliance and in studies examining the nutrient intake of specific populations. Liquid chromatographic methods, primarily with UV absorbance detection, are well established for both fat- and water-soluble measurements, but they do have limitations for certain analytes and may suffer from a lack of specificity in complex matrices. Liquid chromatography-mass spectrometry (LC-MS) provides both sensitivity and specificity for the determination of vitamins in these matrices, and simultaneous analysis of multiple vitamins in a single analysis is often possible. In this work, LC-MS methods were developed for both fat- and water-soluble vitamins and applied to the measurement of these analytes in two NIST Standard Reference Materials. When possible, stable isotope labeled internal standards were employed for quantification.

  11. Phosphine reduced IgG. A new method for 99mTc labeling immunoglobulins

    International Nuclear Information System (INIS)

    Arteaga de Murphy, C.; Melendez-Alafort, L.; Martinez-Rivero, O.; Gomez, E.; Ferro-Flores, G.

    1997-01-01

    A new technetium labeling method for immunoglobulins reduced with tris(2-carboxy-ethyl)phosphine hydrochloride is presented. The Sandoglobulina IgG source was assayed for purity and optimum reagent's concentration and incubation times were determined. It was purified by column chromatography and labelled with Sn 2+ reduced technetium in the presence of MDP. The kit is easy to prepare, labeling efficiency is >(97±1.9)% and stable for 6 hours.The immunoreactivity of the 99 Tc-IgG was verified by electrophoresis and Western blot tests. The IgG retained its structure after both the reducing and labeling processes and it was the only labeled species. (author)

  12. Phosphine reduced IgG. A new method for {sup 99m}Tc labeling immunoglobulins

    Energy Technology Data Exchange (ETDEWEB)

    Arteaga de Murphy, C; Melendez-Alafort, L [Radiofarmacia Departamento de Medicina Nuclear, Instituto Nacional de Nutricion Salvador Zubiran, Mexico (Mexico); Martinez-Rivero, O [Laboratorio de Quimica Organica, Facultad de Quimica, Universidad de la Habana, Habana (Cuba); Gomez, E [Departamento de Fisiologia de la Nutricion, Instituto Nacional de Nutricion Salvador Zubiran, Mexico (Mexico); Ferro-Flores, G [Depeartamento del Reactor y Materiales Radioactivos, Instituto Nacional de Investigaciones Nucleares, Mexico (Mexico)

    1997-09-01

    A new technetium labeling method for immunoglobulins reduced with tris(2-carboxy-ethyl)phosphine hydrochloride is presented. The Sandoglobulina IgG source was assayed for purity and optimum reagent`s concentration and incubation times were determined. It was purified by column chromatography and labelled with Sn{sup 2+} reduced technetium in the presence of MDP. The kit is easy to prepare, labeling efficiency is >(97{+-}1.9)% and stable for 6 hours.The immunoreactivity of the {sup 99}Tc-IgG was verified by electrophoresis and Western blot tests. The IgG retained its structure after both the reducing and labeling processes and it was the only labeled species. (author). 11 refs.

  13. Experimental design-based isotope-dilution SPME-GC/MS method development for the analysis of smoke flavouring products.

    Science.gov (United States)

    Giri, Anupam; Zelinkova, Zuzana; Wenzl, Thomas

    2017-12-01

    For the implementation of Regulation (EC) No 2065/2003 related to smoke flavourings used or intended for use in or on foods a method based on solid-phase micro extraction (SPME) GC/MS was developed for the characterisation of liquid smoke products. A statistically based experimental design (DoE) was used for method optimisation. The best general conditions to quantitatively analyse the liquid smoke compounds were obtained with a polydimethylsiloxane/divinylbenzene (PDMS/DVB) fibre, 60°C extraction temperature, 30 min extraction time, 250°C desorption temperature, 180 s desorption time, 15 s agitation time, and 250 rpm agitation speed. Under the optimised conditions, 119 wood pyrolysis products including furan/pyran derivatives, phenols, guaiacol, syringol, benzenediol, and their derivatives, cyclic ketones, and several other heterocyclic compounds were identified. The proposed method was repeatable (RSD% <5) and the calibration functions were linear for all compounds under study. Nine isotopically labelled internal standards were used for improving quantification of analytes by compensating matrix effects that might affect headspace equilibrium and extractability of compounds. The optimised isotope dilution SPME-GC/MS based analytical method proved to be fit for purpose, allowing the rapid identification and quantification of volatile compounds in liquid smoke flavourings.

  14. Formic acid hydrolysis/liquid chromatography isotope dilution mass spectrometry: An accurate method for large DNA quantification.

    Science.gov (United States)

    Shibayama, Sachie; Fujii, Shin-Ichiro; Inagaki, Kazumi; Yamazaki, Taichi; Takatsu, Akiko

    2016-10-14

    Liquid chromatography-isotope dilution mass spectrometry (LC-IDMS) with formic acid hydrolysis was established for the accurate quantification of λDNA. The over-decomposition of nucleobases in formic acid hydrolysis was restricted by optimizing the reaction temperature and the reaction time, and accurately corrected by using deoxynucleotides (dNMPs) and isotope-labeled dNMPs as the calibrator and the internal standard, respectively. The present method could quantify λDNA with an expanded uncertainty of 4.6% using 10fmol of λDNA. The analytical results obtained with the present method were validated by comparing with the results of phosphate-base quantification by inductively coupled plasma-mass spectrometry (ICP-MS). The results showed good agreement with each other. We conclude that the formic acid hydrolysis/LC-IDMS method can quantify λDNA accurately and is promising as the primary method for the certification of DNA as reference material. Copyright © 2016 Elsevier B.V. All rights reserved.

  15. Assessment of methods for analyzing gaseous mixtures of hydrogen isotopes and helium

    International Nuclear Information System (INIS)

    Attalla, A.; Bishop, C.T.; Bohl, D.R.; Buxton, T.L.; Sprague, R.E.; Warner, D.K.

    1976-01-01

    Mass spectrographic methods have served well in the past to analyze gaseous mixtures of the hydrogen isotopes. Alternate methods of analyses are reviewed which offer wider ranges and variety of isotopic determinations. This report describes possible improvements of the mass spectrographic determinations, gas chromatography, anti-Stokes Raman spectroscopy, microwave-induced optical emission spectroscopy, and methods of measuring tritium using radiation detection devices. Precision, accuracy, limitations, and costs are included for some of the methods mentioned. Costs range from $70,000 for the anti-Stokes Raman spectroscopy equipment, which can determine hydrogen isotopes but not helium, to less than $10,000 for the gas chromatographic equipment, which can determine hydrogen isotopes and helium with precision and accuracy comparable to those of the mass spectrometer

  16. Transport of Indole-3-Butyric Acid and Indole-3-Acetic Acid in Arabidopsis Hypocotyls Using Stable Isotope Labeling1[C][W][OA

    Science.gov (United States)

    Liu, Xing; Barkawi, Lana; Gardner, Gary; Cohen, Jerry D.

    2012-01-01

    The polar transport of the natural auxins indole-3-butyric acid (IBA) and indole-3-acetic acid (IAA) has been described in Arabidopsis (Arabidopsis thaliana) hypocotyls using radioactive tracers. Because radioactive assays alone cannot distinguish IBA from its metabolites, the detected transport from applied [3H]IBA may have resulted from the transport of IBA metabolites, including IAA. To test this hypothesis, we used a mass spectrometry-based method to quantify the transport of IBA in Arabidopsis hypocotyls by following the movement of [13C1]IBA and the [13C1]IAA derived from [13C1]IBA. We also assayed [13C6]IAA transport in a parallel control experiment. We found that the amount of transported [13C1]IBA was dramatically lower than [13C6]IAA, and the IBA transport was not reduced by the auxin transport inhibitor N-1-naphthylphthalamic acid. Significant amounts of the applied [13C1]IBA were converted to [13C1]IAA during transport, but [13C1]IBA transport was independent of IBA-to-IAA conversion. We also found that most of the [13C1]IBA was converted to ester-linked [13C1]IBA at the apical end of hypocotyls, and ester-linked [13C1]IBA was also found in the basal end at a level higher than free [13C1]IBA. In contrast, most of the [13C6]IAA was converted to amide-linked [13C6]IAA at the apical end of hypocotyls, but very little conjugated [13C6]IAA was found in the basal end. Our results demonstrate that the polar transport of IBA is much lower than IAA in Arabidopsis hypocotyls, and the transport mechanism is distinct from IAA transport. These experiments also establish a method for quantifying the movement of small molecules in plants using stable isotope labeling. PMID:22323783

  17. Adsorption methods for hydrogen isotope storage on zeolitic sieves

    International Nuclear Information System (INIS)

    Cristescu, Ioana; Cristescu, Ion; Vasut, F.; Brad, S.; Lazar, A.

    2001-01-01

    For hydrogen isotope separation, adsorption molecular sieves and active carbon were used. Adsorption process proceeds at liquid nitrogen and liquid hydrogen temperatures. Commercial zeolites have the same proprieties with natural zeolites, but they have a regular pore structure. They also have affinity for molecules of different size with defined shapes. Experimental results obtained at liquid nitrogen temperature (77.4 K) and liquid hydrogen revealed the efficient behaviour of the active carbon and zeolitic sieves for hydrogen isotopes temporary storage. We study adsorption of the synthetic zeolites in a wide range of temperatures and pressures and we used the molecular sieves 4A, 5A and active carbon. The 4A and 5A zeolites have a tridimensional structure with 11.4 A diameter. When the hydration water is eliminated, the material keeps a porous structure. The porous volume represents 45% from the zeolite mass for 4A and 5A sieves. The activation temperature of the zeolite and the carbon is very important for obtaining a high adsorption capacity. If the temperature used for activation is low, the structural water will be not eliminated and the adsorption capacity will be low. The excessive temperature will destroy the porous structure. The adsorption capacity for the hydrogen isotopes was calculated with the relation: A = V ads /m (cm 3 /g). The adsorption capacity and efficiency for the adsorbent materials, are given. Physical adsorption process of the hydrogen isotopes was carried out at liquid nitrogen temperature. The flux gas used in the adsorption system is composed of dry deuterium and protium. This mixture is cooled in liquid nitrogen and then is passed to the adsorbent getter at the same temperature (77.4 K). The gas flux in the adsorbent getter is 5 and 72 l/h (which correspond to 0.008 and 0.134 discharge velocity, respectively). (authors)

  18. Stable isotopes and elasmobranchs: tissue types, methods, applications and assumptions.

    Science.gov (United States)

    Hussey, N E; MacNeil, M A; Olin, J A; McMeans, B C; Kinney, M J; Chapman, D D; Fisk, A T

    2012-04-01

    Stable-isotope analysis (SIA) can act as a powerful ecological tracer with which to examine diet, trophic position and movement, as well as more complex questions pertaining to community dynamics and feeding strategies or behaviour among aquatic organisms. With major advances in the understanding of the methodological approaches and assumptions of SIA through dedicated experimental work in the broader literature coupled with the inherent difficulty of studying typically large, highly mobile marine predators, SIA is increasingly being used to investigate the ecology of elasmobranchs (sharks, skates and rays). Here, the current state of SIA in elasmobranchs is reviewed, focusing on available tissues for analysis, methodological issues relating to the effects of lipid extraction and urea, the experimental dynamics of isotopic incorporation, diet-tissue discrimination factors, estimating trophic position, diet and mixing models and individual specialization and niche-width analyses. These areas are discussed in terms of assumptions made when applying SIA to the study of elasmobranch ecology and the requirement that investigators standardize analytical approaches. Recommendations are made for future SIA experimental work that would improve understanding of stable-isotope dynamics and advance their application in the study of sharks, skates and rays. © 2012 The Authors. Journal of Fish Biology © 2012 The Fisheries Society of the British Isles.

  19. Method of concentrating and separating lithium isotopes by laser

    International Nuclear Information System (INIS)

    Yamashita, Mikio; Kashiwagi, Hiroshi.

    1976-01-01

    Purpose: To eliminate the need for repeating separating operation in many stages and permit concentrated lithium of high purity to be obtained in a short period of time. Constitution: Lithium atom vapor is irradiated by a laser of wavelengths resonant to 6 Li or 7 Li absorption spectra present in the neighborhood of 6,707.84 A or 3,232.61 A (chromatic laser being used for oscillation in the neighborhood of 6,707.84 A and ultraviolet laser used for oscillation in the neighborhood of 6,707.84 A) for selectively exciting 6 Li or 7 Li alone. Then, ionization is brought about by using other types of lasers (ultraviolet Ar ion laser being used as the former wavelength laser and visible Ar ion laser as the latter wavelength laser), and only the ionized isotopes are passed through a mass filter and collected by an ion collector, thereby effecting separation of the ionized isotopes from the non-ionized neutral isotopes and their concentration. (Aizawa, K.)

  20. Measurement of Hepatic Protein Fractional Synthetic Rate with Stable Isotope Labeling Technique in Thapsigargin Stressed HepG2 Cells

    Science.gov (United States)

    Song, Juquan; Zhang, Xiao-jun; Boehning, Darren; Brooks, Natasha C.; Herndon, David N.; Jeschke, Marc G.

    2012-01-01

    Severe burn-induced liver damage and dysfunction is associated with endoplasmic reticulum (ER) stress. ER stress has been shown to regulate global protein synthesis. In the current study, we induced ER stress in vitro and estimated the effect of ER stress on hepatic protein synthesis. The aim was two-fold: (1) to establish an in vitro model to isotopically measure hepatic protein synthesis and (2) to evaluate protein fractional synthetic rate (FSR) in response to ER stress. Human hepatocellular carcinoma cells (HepG2) were cultured in medium supplemented with stable isotopes 1,2-13C2-glycine and L-[ring-13C6]phenylalanine. ER stress was induced by exposing the cells to 100 nM of thapsigargin (TG). Cell content was collected from day 0 to 14. Alterations in cytosolic calcium were measured by calcium imaging and ER stress markers were confirmed by Western blotting. The precursor and product enrichments were detected by GC-MS analysis for FSR calculation. We found that the hepatic protein FSR were 0.97±0.02 and 0.99±0.05%/hr calculated from 1,2-13C2-glycine and L-[ring-13C6]phenylalanine, respectively. TG depleted ER calcium stores and induced ER stress by upregulating p-IRE-1 and Bip. FSR dramatically decreased to 0.68±0.03 and 0.60±0.06%/hr in the TG treatment group (pisotope tracer incorporation technique is a useful method for studying the effects of ER stress on hepatic protein synthesis. PMID:22298954

  1. Alcohol Warning Label Awareness and Attention: A Multi-method Study.

    Science.gov (United States)

    Pham, Cuong; Rundle-Thiele, Sharyn; Parkinson, Joy; Li, Shanshi

    2018-01-01

    Evaluation of alcohol warning labels requires careful consideration ensuring that research captures more than awareness given that labels may not be prominent enough to attract attention. This study investigates attention of current in market alcohol warning labels and examines whether attention can be enhanced through theoretically informed design. Attention scores obtained through self-report methods are compared to objective measures (eye-tracking). A multi-method experimental design was used delivering four conditions, namely control, colour, size and colour and size. The first study (n = 559) involved a self-report survey to measure attention. The second study (n = 87) utilized eye-tracking to measure fixation count and duration and time to first fixation. Analysis of Variance (ANOVA) was utilized. Eye-tracking identified that 60% of participants looked at the current in market alcohol warning label while 81% looked at the optimized design (larger and red). In line with observed attention self-reported attention increased for the optimized design. The current study casts doubt on dominant practices (largely self-report), which have been used to evaluate alcohol warning labels. Awareness cannot be used to assess warning label effectiveness in isolation in cases where attention does not occur 100% of the time. Mixed methods permit objective data collection methodologies to be triangulated with surveys to assess warning label effectiveness. Attention should be incorporated as a measure in warning label effectiveness evaluations. Colour and size changes to the existing Australian warning labels aided by theoretically informed design increased attention. © The Author 2017. Medical Council on Alcohol and Oxford University Press. All rights reserved.

  2. Improvement of organic compounds labelling method with the use of thermally activated tritium gas

    International Nuclear Information System (INIS)

    Nejman, L.A.; Smolyakov, V.S.; Antropova, L.P.

    1982-01-01

    Use of a support (various types of papers) is recommended for organic compounds labelling by tritium gas activated at a hot tungsten filament. This improvement increases chemical and radiochemical yields and makes the experiment simpler and faster. Generally labelled triethyloxonium tetra-fluoroborate, ethyl-p-aminobenzoate, p-aminobenzoic acid (Na-salt), A-factor (a natural regulator of streptomycin biosynthesis), decapeptide angiotensin I, phospholipid 1, 2 - dimyristoyl-sn-glycero-3--phosphocholine and E. coli tRNAs have been prepared by this method. Molar radioactivity of the labelled compounds is in the range of 1-200 GBg/mmole [ru

  3. ICT: isotope correction toolbox.

    Science.gov (United States)

    Jungreuthmayer, Christian; Neubauer, Stefan; Mairinger, Teresa; Zanghellini, Jürgen; Hann, Stephan

    2016-01-01

    Isotope tracer experiments are an invaluable technique to analyze and study the metabolism of biological systems. However, isotope labeling experiments are often affected by naturally abundant isotopes especially in cases where mass spectrometric methods make use of derivatization. The correction of these additive interferences--in particular for complex isotopic systems--is numerically challenging and still an emerging field of research. When positional information is generated via collision-induced dissociation, even more complex calculations for isotopic interference correction are necessary. So far, no freely available tools can handle tandem mass spectrometry data. We present isotope correction toolbox, a program that corrects tandem mass isotopomer data from tandem mass spectrometry experiments. Isotope correction toolbox is written in the multi-platform programming language Perl and, therefore, can be used on all commonly available computer platforms. Source code and documentation can be freely obtained under the Artistic License or the GNU General Public License from: https://github.com/jungreuc/isotope_correction_toolbox/ {christian.jungreuthmayer@boku.ac.at,juergen.zanghellini@boku.ac.at} Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  4. Temperature-induced evolution of strain and doping in an isotopically labeled two-dimensional graphene-C-70 fullerene peapod

    Czech Academy of Sciences Publication Activity Database

    Verhagen, Timotheus; Valeš, Václav; Kalbáč, Martin; Vejpravová, Jana

    2017-01-01

    Roč. 75, May (2017), s. 140-145 ISSN 0925-9635 R&D Projects: GA ČR(CZ) GA15-01953S; GA MŠk LL1301 Institutional support: RVO:68378271 ; RVO:61388955 Keywords : two-dimensional peapod * Raman spectroscopy * isotope labelling * topography Subject RIV: BM - Solid Matter Physics ; Magnetism; CF - Physical ; Theoretical Chemistry (UFCH-W) OBOR OECD: Condensed matter physics (including formerly solid state physics, supercond.); Physical chemistry (UFCH-W) Impact factor: 2.561, year: 2016

  5. A comparative study of fat storage quantitation in nematode Caenorhabditis elegans using label and label-free methods.

    Directory of Open Access Journals (Sweden)

    Kelvin Yen

    to the staining of fat stores, but rather the sequestration of dyes in lysosome-related organelles. In contrast, fixative staining methods provide reproducible data but are prone to errors due to the interference of autofluorescent species and the non-specific staining of cellular structures other than fat stores. Importantly, both growth conditions and developmental stage should be considered when comparing methods of C. elegans lipid storage. Taken together, we confirm that CARS microscopy provides a direct, non-invasive, and label-free means to quantitatively analyze fat storage in living C. elegans.

  6. A SIMPLE FLUORESCENT LABELING METHOD FOR STUDIES OF PROTEIN OXIDATION, PROTEIN MODIFICATION, AND PROTEOLYSIS

    Science.gov (United States)

    Pickering, Andrew. M.; Davies, Kelvin. J. A.

    2014-01-01

    Proteins are sensitive to oxidation, and oxidized proteins are excellent substrates for degradation by proteolytic enzymes such as the Proteasome and the mitochondrial Lon protease. Protein labeling is required for studies of protein turnover. Unfortunately, most labeling techniques involve 3H or 14C methylation which is expensive, exposes researchers to radioactivity, generates large amounts of radioactive waste, and allows only single-point assays because samples require acid-precipitation. Alternative labeling methods, have largely proven unsuitable, either because the probe itself is modified by the oxidant(s) being studied, or because the alternative labeling techniques are too complex or too costly for routine use. What is needed is a simple, quick, and cheap labeling technique that uses a non-radioactive marker, that binds strongly to proteins, is resistant to oxidative modification, and emits a strong signal. We have devised a new reductive method for labeling free carboxyl groups of proteins with the small fluorophore 7-amino-4-methycoumarin (AMC). When bound to target proteins, AMC fluoresces very weakly but when AMC is released by proteinases, proteases, or peptidases, it fluoresces strongly. Thus, without acid-precipitation, the proteolysis of any target protein can be studied continuously, in multiwell plates. In direct comparisons, 3H-labeled proteins and AMC-labeled proteins exhibited essentially identical degradation patterns during incubation with trypsin, cell extracts, and purified proteasome. AMC-labeled proteins are well-suited to study increased proteolytic susceptibility following protein modification, since the AMC-protein bond is resistant to oxidizing agents such as hydrogen peroxide and peroxynitrite, and is stable over time and to extremes of pH, temperature (even boiling), freeze-thawing, mercaptoethanol, and methanol. PMID:21988844

  7. Mass spectrometric measurements of norepinephrine synthesis in man from infusion of stable isotope-labelled L-threo-3,4-dihydroxyphenylserine

    International Nuclear Information System (INIS)

    Suzuki, T.; Sakoda, S.; Ueji, M.; Kishimoto, S.

    1985-01-01

    The kinetics of stable isotope-labelled L-threo-3,4-dihydroxyphenylserine (L-threo-DOPS), an immediate precursor of (-)-norepinephrine, was studied to investigate the pharmacologic mechanism of its therapeutic effect on orthostatic hypotension in familial amyloid polyneuropathy (FAP) and on akinesia and freezing in parkinsonism. [ 13 C,D]-L-threo-DOPS was synthesized, and 100 mg of the compound was infused for 2 h into two normal subjects, two FAP patients and two patients with the degenerative diseases of the central nervous system. Labelled and endogenous norepinephrine in urine and plasma was assayed simultaneously by gas chromatography/mass spectrometry. The results indicate that the increase in norepinephrine in biological fluids after administration of L-threo-DOPS is attributable mostly to norepinephrine derived from L-threo-DOPS, not to pre-formed endogenous norepinephrine released by L-threo-DOPS

  8. Inverse methods for estimating primary input signals from time-averaged isotope profiles

    Science.gov (United States)

    Passey, Benjamin H.; Cerling, Thure E.; Schuster, Gerard T.; Robinson, Todd F.; Roeder, Beverly L.; Krueger, Stephen K.

    2005-08-01

    Mammalian teeth are invaluable archives of ancient seasonality because they record along their growth axes an isotopic record of temporal change in environment, plant diet, and animal behavior. A major problem with the intra-tooth method is that intra-tooth isotope profiles can be extremely time-averaged compared to the actual pattern of isotopic variation experienced by the animal during tooth formation. This time-averaging is a result of the temporal and spatial characteristics of amelogenesis (tooth enamel formation), and also results from laboratory sampling. This paper develops and evaluates an inverse method for reconstructing original input signals from time-averaged intra-tooth isotope profiles. The method requires that the temporal and spatial patterns of amelogenesis are known for the specific tooth and uses a minimum length solution of the linear system Am = d, where d is the measured isotopic profile, A is a matrix describing temporal and spatial averaging during amelogenesis and sampling, and m is the input vector that is sought. Accuracy is dependent on several factors, including the total measurement error and the isotopic structure of the measured profile. The method is shown to accurately reconstruct known input signals for synthetic tooth enamel profiles and the known input signal for a rabbit that underwent controlled dietary changes. Application to carbon isotope profiles of modern hippopotamus canines reveals detailed dietary histories that are not apparent from the measured data alone. Inverse methods show promise as an effective means of dealing with the time-averaging problem in studies of intra-tooth isotopic variation.

  9. Isotope methods in water resources assessment and environmental management

    International Nuclear Information System (INIS)

    Araguas-Araguas, L.

    1996-01-01

    Availability of water and protection of water resources have become top environmental issues in many countries. Governments are forced to issue strict guidelines to protect the environment and create agencies to pursue these aspects as well as enforce such regulations. The supply of good-quality water from rivers and lakes is becoming a costly and complex problem for many institutes responsible for water supply. Because of the high pollution levels in surface waters, ground water is the main source of drinking water in many countries. It is estimated that 1.5 billion people world-wide depend on it for drinking water. Since ground water cannot be directly measured, and despite its importance for drinking purposes there is not enough public concern about its protection. In other cases, it is found that the exploited ground water is not a renewable resource. In many countries in arid and semi-arid regions, fossil ground water is being tapped for extensive agricultural development, but such extraction depletes the reserves, in the same way as an oil reservoir. The availability of correct information, before decisions are taken will lead to improved management of water resources, distributing the available resources for different uses according to their quality, and ultimately, to manage the resource. Nuclear science has developed a series of methodologies based on the use of naturally-occurring isotopes and artificial tracers to study the processes involved in the occurrence and circulation of water. The discipline called 'Isotope Hydrology' provides a deep insight into many parts of the water cycle; from the evaporation over the ocean or the continents, to the formation of surface runoff and ground water and in the discharge of aquifer systems into the ocean. Isotope hydrology, as a scientific and applied discipline in earth sciences, was created during the late 1950s and early 1960s, beyond the classical hydrological science. In these early stages, new methodologies

  10. Extraction method for the determination of inorganic iodides in Rose Bengal labelled with 131I

    International Nuclear Information System (INIS)

    Lengyel, J.; Krtil, J.; Vecernik, J.

    1982-01-01

    An extraction method for the determination of inorganic iodides in Rose Bengal preparations labelled with 131 I is described. The method is based on the quantitative extraction of Rose Bengal into chloroform from acidic medium while the inorganic iodides remain in the aqueous phase. The method is simple, rapid, and reproducible. (author)

  11. Method of deuterium isotope separation using ethylene and ethylene dichloride

    International Nuclear Information System (INIS)

    Benson, S.W.

    1982-01-01

    Compounds enriched in deuterium may be obtained from ethylene, vinyl chloride, 1,2-dichloroethane, or propylene by laser isotope separation. Normal molecules of these organic compounds are exposed to infrared laser radiation of a suitable wavelength. Substantially all of the deuterium-containing molecules exposed to the laser can be selectively dissociated and the deuterium-containing products separated from the starting material and other reaction products. The deuterium-containing molecules can be burned to form water with an enriched deuterium content, or pyrolized to form hydrogen gas enriched in deuterium

  12. Application and validation of isotope dilution method (IDM) for predicting bioavailability of hydrophobic organic contaminants in soil.

    Science.gov (United States)

    Wang, Jie; Taylor, Allison; Schlenk, Daniel; Gan, Jay

    2018-05-01

    Risk assessment of hydrophobic organic contaminants (HOCs) using the total chemical concentration following exhaustive extraction may overestimate the actual availability of HOCs to non-target organisms. Existing methods for estimating HOC bioavailability in soil have various operational limitations. In this study, we explored the application of isotope dilution method (IDM) to quantify the accessible fraction (E) of DDTs and PCBs in both historically-contaminated and freshly-spiked soils. After addition of 13 C or deuterated analogues to a soil sample, the phase distribution of isotope-labeled and native chemicals reached an apparent equilibrium within 48 h of mixing. The derived E values in the three soils ranged from 0.19 to 0.82, depending on the soil properties and also the contact time of HOCs (i.e., aging). The isotope dilution method consistently predicted greater accumulation into earthworm (Eisenia fetida) than that by polyethylene (PE) or solid phase microextraction (SPME) sampler, likely because desorption in the gut enhanced bioavailability of soil-borne HOCs. A highly significant linear regression (R 2  = 0.91) was found between IDM and 24-h Tenax desorption, with a slope statistically identical to 1. The IDM-derived accessible concentration (C e ) was further shown to accurately predict tissue residues in earthworm exposed in the same soils. Given the relatively short duration and simple steps, IDM has the potential to be readily adopted for measuring HOC bioaccessibility in soil and for improving risk assessment and evaluation of remediation efficiency. Copyright © 2018 Elsevier Ltd. All rights reserved.

  13. Introducing AAA-MS, a rapid and sensitive method for amino acid analysis using isotope dilution and high-resolution mass spectrometry.

    Science.gov (United States)

    Louwagie, Mathilde; Kieffer-Jaquinod, Sylvie; Dupierris, Véronique; Couté, Yohann; Bruley, Christophe; Garin, Jérôme; Dupuis, Alain; Jaquinod, Michel; Brun, Virginie

    2012-07-06

    Accurate quantification of pure peptides and proteins is essential for biotechnology, clinical chemistry, proteomics, and systems biology. The reference method to quantify peptides and proteins is amino acid analysis (AAA). This consists of an acidic hydrolysis followed by chromatographic separation and spectrophotometric detection of amino acids. Although widely used, this method displays some limitations, in particular the need for large amounts of starting material. Driven by the need to quantify isotope-dilution standards used for absolute quantitative proteomics, particularly stable isotope-labeled (SIL) peptides and PSAQ proteins, we developed a new AAA assay (AAA-MS). This method requires neither derivatization nor chromatographic separation of amino acids. It is based on rapid microwave-assisted acidic hydrolysis followed by high-resolution mass spectrometry analysis of amino acids. Quantification is performed by comparing MS signals from labeled amino acids (SIL peptide- and PSAQ-derived) with those of unlabeled amino acids originating from co-hydrolyzed NIST standard reference materials. For both SIL peptides and PSAQ standards, AAA-MS quantification results were consistent with classical AAA measurements. Compared to AAA assay, AAA-MS was much faster and was 100-fold more sensitive for peptide and protein quantification. Finally, thanks to the development of a labeled protein standard, we also extended AAA-MS analysis to the quantification of unlabeled proteins.

  14. The outline of the processes for lithium isotope separation by ion exchange method

    International Nuclear Information System (INIS)

    Fujine, Sachio; Saito, Keiichiro; Naruse, Yuji; Shiba, Koreyuki; Kosuge, Masao; Itoi, Toshiaki; Kitsukawa, Tomohiko.

    1981-10-01

    A plant of lithium isotope separation by displacement chromatography is preliminary designed. The construction expenses of a 100 kg 7 Li/year plant and the unit cost of separation are estimated on the basis of the data taken from the literature, and the feasibility is studied. Experimental equipment of continuous displacement chromatography is set up and is tested with the stable automatic operation. These results indicate that the ion exchange method is promising for industrial lithium isotope separation. (author)

  15. Ground-state properties of axially deformed Sr isotopes in Skyrme-Hartree-Fock-Bogolyubov method

    International Nuclear Information System (INIS)

    Yilmaz, A.H.; Bayram, T.; Demirci, M.; Engin, B.; Bayram, T.

    2010-01-01

    Binding energies, the mean-square nuclear radii, neutron radii, quadrupole moments and deformation parameters to axially deformed Strontium isotopes were evaluated using Hartree-Fock-Bogolyubov method. Shape coexistence was also discussed. The results were compared with experimental data and some estimates obtained within some nuclear models. The calculations were performed for SIy4 set of Skyrme forces and for wide range of the neutron numbers of Sr isotopes

  16. Graphite Isotope Ratio Method Development Report: Irradiation Test Demonstration of Uranium as a Low Fluence Indicator

    International Nuclear Information System (INIS)

    Reid, B.D.; Gerlach, D.C.; Love, E.F.; McNeece, J.P.; Livingston, J.V.; Greenwood, L.R.; Petersen, S.L.; Morgan, W.C.

    1999-01-01

    This report describes an irradiation test designed to investigate the suitability of uranium as a graphite isotope ratio method (GIRM) low fluence indicator. GIRM is a demonstrated concept that gives a graphite-moderated reactor's lifetime production based on measuring changes in the isotopic ratio of elements known to exist in trace quantities within reactor-grade graphite. Appendix I of this report provides a tutorial on the GIRM concept

  17. Innovative method for ultra-sensitive measurement of krypton isotopes

    International Nuclear Information System (INIS)

    Lavielle, B.; Gilabert, E.; Thomas, B.; Rebeix, R.; Canchel, G.; Moulin, C.; Topin, S.; Pointurier, F.

    2015-01-01

    Kr 81 (T 1/2 = 2.29*10 5 y), that is produced in the atmosphere by cosmic rays and Kr 85 (T 1/2 = 10.77 y), that is produced in the fission of nuclear fuels, are considered as the right chronometer elements for the dating of underground waters, polar ice, of for tracking civil and military nuclear activities. The use of Kr 81 and Kr 85 implies the development of extracting lines and detection devices with so high accuracy that only a few thousands of krypton atoms can be detected. The approach developed aims at detecting Kr 81 and Kr 85 in 1 liter of air and 20 liters of water. 3 steps are necessary. The first step consists in separating and purifying the krypton. The extraction of gases from water is made by helium bubbling. Chemically active gases like N 2 , CO 2 , CH 4 and O 2 are eliminated by chemical traps while Ar si separated by cryogenics. The second step involves a double focus mass spectrometer designed to perform an important enrichment in Kr 81 and Kr 85 . The last step is the high-accuracy measurement of krypton isotopes performed with the new tool named FAKIR (Facility for Analyzing Krypton Isotopic Ratios) that is based on UV laser ionization and on the mass-discrimination of the ions through their time of flight

  18. Selenium-75-labelled foliate compounds

    International Nuclear Information System (INIS)

    1974-01-01

    A saturation method to analyze a foliate is presented; it uses competitive reaction of the compound to be measured and of a radioactive-labelled version of this compound with a reagent specific to this compound present in insufficient quantity to combine with the whole of the compound and its labelled version, separation of the bound compound from its non-bound homologue and measurement of the radioactivity concentration in the bound compound, the non-bound compound or both. The radioactive isotope used in the labelled foliate is selenium 75 [fr

  19. Novel methods for estimating 3D distributions of radioactive isotopes in materials

    Energy Technology Data Exchange (ETDEWEB)

    Iwamoto, Y., E-mail: y.iwamoto0805@ruri.waseda.jp [Research Institute for Science and Engineering, Waseda University, Shinjuku, Tokyo 169-8555 (Japan); Kataoka, J.; Kishimoto, A.; Nishiyama, T.; Taya, T.; Okochi, H.; Ogata, H. [Research Institute for Science and Engineering, Waseda University, Shinjuku, Tokyo 169-8555 (Japan); Yamamoto, S. [Nagoya University Graduate School of Medicine, 1-1-20, Daikominami, Higashi-ku, Nagoya-shi, Aichi 461-8673 (Japan)

    2016-09-21

    In recent years, various gamma-ray visualization techniques, or gamma cameras, have been proposed. These techniques are extremely effective for identifying “hot spots” or regions where radioactive isotopes are accumulated. Examples of such would be nuclear-disaster-affected areas such as Fukushima or the vicinity of nuclear reactors. However, the images acquired with a gamma camera do not include distance information between radioactive isotopes and the camera, and hence are “degenerated” in the direction of the isotopes. Moreover, depth information in the images is lost when the isotopes are embedded in materials, such as water, sand, and concrete. Here, we propose two methods of obtaining depth information of radioactive isotopes embedded in materials by comparing (1) their spectra and (2) images of incident gamma rays scattered by the materials and direct gamma rays. In the first method, the spectra of radioactive isotopes and the ratios of scattered to direct gamma rays are obtained. We verify experimentally that the ratio increases with increasing depth, as predicted by simulations. Although the method using energy spectra has been studied for a long time, an advantage of our method is the use of low-energy (50–150 keV) photons as scattered gamma rays. In the second method, the spatial extent of images obtained for direct and scattered gamma rays is compared. By performing detailed Monte Carlo simulations using Geant4, we verify that the spatial extent of the position where gamma rays are scattered increases with increasing depth. To demonstrate this, we are developing various gamma cameras to compare low-energy (scattered) gamma-ray images with fully photo-absorbed gamma-ray images. We also demonstrate that the 3D reconstruction of isotopes/hotspots is possible with our proposed methods. These methods have potential applications in the medical fields, and in severe environments such as the nuclear-disaster-affected areas in Fukushima.

  20. Novel methods for estimating 3D distributions of radioactive isotopes in materials

    Science.gov (United States)

    Iwamoto, Y.; Kataoka, J.; Kishimoto, A.; Nishiyama, T.; Taya, T.; Okochi, H.; Ogata, H.; Yamamoto, S.

    2016-09-01

    In recent years, various gamma-ray visualization techniques, or gamma cameras, have been proposed. These techniques are extremely effective for identifying "hot spots" or regions where radioactive isotopes are accumulated. Examples of such would be nuclear-disaster-affected areas such as Fukushima or the vicinity of nuclear reactors. However, the images acquired with a gamma camera do not include distance information between radioactive isotopes and the camera, and hence are "degenerated" in the direction of the isotopes. Moreover, depth information in the images is lost when the isotopes are embedded in materials, such as water, sand, and concrete. Here, we propose two methods of obtaining depth information of radioactive isotopes embedded in materials by comparing (1) their spectra and (2) images of incident gamma rays scattered by the materials and direct gamma rays. In the first method, the spectra of radioactive isotopes and the ratios of scattered to direct gamma rays are obtained. We verify experimentally that the ratio increases with increasing depth, as predicted by simulations. Although the method using energy spectra has been studied for a long time, an advantage of our method is the use of low-energy (50-150 keV) photons as scattered gamma rays. In the second method, the spatial extent of images obtained for direct and scattered gamma rays is compared. By performing detailed Monte Carlo simulations using Geant4, we verify that the spatial extent of the position where gamma rays are scattered increases with increasing depth. To demonstrate this, we are developing various gamma cameras to compare low-energy (scattered) gamma-ray images with fully photo-absorbed gamma-ray images. We also demonstrate that the 3D reconstruction of isotopes/hotspots is possible with our proposed methods. These methods have potential applications in the medical fields, and in severe environments such as the nuclear-disaster-affected areas in Fukushima.

  1. Differential quantitative proteomics of Porphyromonas gingivalis by linear ion trap mass spectrometry: Non-label methods comparison, q-values and LOWESS curve fitting

    Science.gov (United States)

    Xia, Qiangwei; Wang, Tiansong; Park, Yoonsuk; Lamont, Richard J.; Hackett, Murray

    2007-01-01

    Differential analysis of whole cell proteomes by mass spectrometry has largely been applied using various forms of stable isotope labeling. While metabolic stable isotope labeling has been the method of choice, it is often not possible to apply such an approach. Four different label free ways of calculating expression ratios in a classic "two-state" experiment are compared: signal intensity at the peptide level, signal intensity at the protein level, spectral counting at the peptide level, and spectral counting at the protein level. The quantitative data were mined from a dataset of 1245 qualitatively identified proteins, about 56% of the protein encoding open reading frames from Porphyromonas gingivalis, a Gram-negative intracellular pathogen being studied under extracellular and intracellular conditions. Two different control populations were compared against P. gingivalis internalized within a model human target cell line. The q-value statistic, a measure of false discovery rate previously applied to transcription microarrays, was applied to proteomics data. For spectral counting, the most logically consistent estimate of random error came from applying the locally weighted scatter plot smoothing procedure (LOWESS) to the most extreme ratios generated from a control technical replicate, thus setting upper and lower bounds for the region of experimentally observed random error.

  2. Development of a new method for hydrogen isotope analysis of trace hydrocarbons in natural gas samples

    Directory of Open Access Journals (Sweden)

    Xibin Wang

    2016-12-01

    Full Text Available A new method had been developed for the analysis of hydrogen isotopic composition of trace hydrocarbons in natural gas samples by using solid phase microextraction (SPME combined with gas chromatography-isotope ratio mass spectrometry (GC/IRMS. In this study, the SPME technique had been initially introduced to achieve the enrichment of trace content of hydrocarbons with low abundance and coupled to GC/IRMS for hydrogen isotopic analysis. The main parameters, including the equilibration time, extraction temperature, and the fiber type, were systematically optimized. The results not only demonstrated that high extraction yield was true but also shows that the hydrogen isotopic fractionation was not observed during the extraction process, when the SPME device fitted with polydimethylsiloxane/divinylbenzene/carbon molecular sieve (PDMS/DVB/CAR fiber. The applications of SPME-GC/IRMS method were evaluated by using natural gas samples collected from different sedimentary basins; the standard deviation (SD was better than 4‰ for reproducible measurements; and also, the hydrogen isotope values from C1 to C9 can be obtained with satisfying repeatability. The SPME-GC/IRMS method fitted with PDMS/DVB/CAR fiber is well suited for the preconcentration of trace hydrocarbons, and provides a reliable hydrogen isotopic analysis for trace hydrocarbons in natural gas samples.

  3. A rapid and convenient method for specific 11C-labelling of synthetic polypeptides containing methionine

    International Nuclear Information System (INIS)

    Laengstroem, B.; Sjoeberg, S.; Ragnarsson, U.

    1981-01-01

    11 C-labelling of methionine residues in a synthetic peptide via the preparation of the corresponding protected, pure homocysteine peptide has been investigated. Complete deprotection of the peptide and specific methylation of the homocysteine residue can be performed in one step in liquid ammonia. As a first application of this method the synthesis of the tripeptide, Z-Gly-L-Hcy(Bzl)-Gly-O-Bzl, and its conversion to Gly-Met-Gly and the corresponding labelled Gly-([ 11 C]-methyl)-Met-Gly, is reported. Starting with the protected peptide the labelling was performed in 20 +- 5 min (starting with 11 CO 2 ), yielding the labelled peptide in 92 +- 5 % radiochemical yield. Analyses and preparative LC can be performed within 6 min. (author)

  4. Chemical method of labelling proteins with the radionuclides of technetium at physiological condition

    International Nuclear Information System (INIS)

    Wong, D.W.

    1983-01-01

    A novel rapid chemical method of labeling plasma proteins, other compounds and/or substances containing protein with radionuclides of technetium such as sup(95m)Tc, sup(99m)Tc or sup(99)Tc at physiologic pH 7.4 condition, producing a sterile non-pyrogenic radioactive tracer material suitable for biological and medical uses. These radiolabeled protein substances are not denatured by the labeling process but retain their natural physiological and immunological properties. This novel labeling technique provides a simple and rapid means of labeling plasma proteins such as human serum albumin, fibrinogen, antibodies, hormones and enzymes with sup(95m)Tc or sup(99m)Tc for scintigraphic imaging which may allow visualization of thrombi, emboli, myocardial infarcts, infectious lesions or tumors

  5. Accurate Determination of Leucine and Valine Side-chain Conformations using U-[15N/13C/2H]/[1H-(methine/methyl)-Leu/Val] Isotope Labeling, NOE Pattern Recognition, and Methine Cγ-Hγ/Cβ-Hβ Residual Dipolar Couplings

    International Nuclear Information System (INIS)

    Tang, Chun; Iwahara, Junji; Clore, G. Marius

    2005-01-01

    An isotope labeling scheme is described in which specific protonation of methine and methyl protons of leucine and valine is obtained on a 15 N/ 13 C labeled background with uniform deuteration of all other non-exchangeable protons. The presence of a protonated methine group has little effect on the favorable relaxation properties of the methyl protons of Leu and Val. This labeling scheme permits the rotameric state of leucine side-chains to be readily determined by simple inspection of the pattern of Hγ(i)-H N (i) and Hγ(i)-H N (i+1) NOEs in a 3D 15 N-separated NOE spectrum free of complications arising from spectral overlap and spin-diffusion. In addition, one-bond residual dipolar couplings for the methine 13 C- 1 H bond vectors of Leu and Val can be accurately determined from an intensity J-modulated constant-time HCCH-COSY experiment and used to accurately orient the side-chains of Leu and Val. Incorporation of these data into structure refinement improves the accuracy with which the conformations of Leu and Val side-chains can be established. This is important to ensure optimal packing both within the protein core and at intermolecular interfaces. The impact of the method on protein structure determination is illustrated by application to enzyme IIA Chitobiose , a 34 kDa homotrimeric phosphotransferase protein

  6. Analysis method for beta-gamma coincidence spectra from radio-xenon isotopes

    International Nuclear Information System (INIS)

    Yang Wenjing; Yin Jingpeng; Huang Xiongliang; Cheng Zhiwei; Shen Maoquan; Zhang Yang

    2012-01-01

    Radio-xenon isotopes monitoring is one important method for the verification of CTBT, what includes the measurement methods of HPGe γ spectrometer and β-γ coincidence. The article describes the analytic flowchart and method of three-dimensional beta-gamma coincidence spectra from β-γ systems, and analyses in detail the principles and methods of the regions of interest of coincidence spectra and subtracting the interference, finally gives the formula of radioactivity of Xenon isotopes and minimum detectable concentrations. Studying on the principles of three-dimensional beta-gamma coincidence spectra, which can supply the foundation for designing the software of β-γ coincidence systems. (authors)

  7. Assessment of phyto-available cadmium in soils using isotopic methods

    International Nuclear Information System (INIS)

    Gerard, Emilie

    2000-01-01

    The quantification of phyto-available Cd in soils is necessary to determine the transfer risk of this toxic element to plants. Isotopic methods (isotopic exchange kinetics (IEK), isotopic dilution) were used to characterize the phyto-available Cd and, practically, to select chemical methods for determining available Cd to plants. Rye grass (Lolium perenne L.), lettuce (Lactuca saliva L.) and the Cd hyper-accumulator Thlaspi caerulescens J. Presl. and C. Presl. were selected as test plants because of their wide range of Cd uptake ability. The chosen soils had different pH and displayed a Cd-contamination gradient due the atmospheric deposition of industrial particles. In the acidic soil, plants had access to the same metal pool, the one that was isotopically exchangeable with Cd 2+ . This was also the case for rye grass on the calcareous Soils but there, lettuce and T. caerulescens accessed a bigger and non-isotopically exchangeable pool that accounted for 16 to 52 % of the plant available Cd. Most of the Cd was isotopically exchangeable in a short time in the acidic soil (87 % within 21 days). In the calcareous soils two pools were identified, one pool was isotopically exchangeable in a very short time, and the other one Was not exchangeable after 21 days (21 to 10 % of the total Cd). Actually, the quantities of labile cd in the industrial particles which had contaminated these soils were very small. IEK methods and the measurement of the isotopic composition of Cd 2+ in soil solutions allowed for the estimation of the phyto-available Cd in soils, but cannot he commonly used. Most of the chemical extractants studied such as CaCl 2 or DTP A are suitable to assess the phyto-available Cd. However, on calcareous soils, the choice of extractant needs to sometimes be modified in relation to the rate of Cd plant absorption. (author) [fr

  8. A rapid method for the preparation of 99Tcm hexametazime-labelled leucocytes

    International Nuclear Information System (INIS)

    Solanki, K.K.; Mather, S.J.; Janabi, M.A.; Britton, K.E.

    1988-01-01

    99 Tc m (±)-hexamethylpropyleneamineoxime (HMPAO) ( 99 Tc m Hexametazime) has been recently reported as an alternative for labelling leucocytes. This technique has been modified to give a simpler routine in-house labelling technique. It has thr