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1.
Replacement of unlabeled -butyrobetaine with -[2,3,4-2H6]butyrobetaine has a profound effect on the stoichiometry between decarboxylation of 2-oxoglutarate and hydroxylation in the reaction catalyzed by human -butyrobetaine hydroxylase. The ratios between decarboxylation and hydroxylation are 1.16 with Unlabeled and 7.48 with deuterated -butyrobetaine as substrate. From these ratios an internal isotope effect of 41 has been calculated. DV in the overall reaction measured as 2- oxoglutarate decarboxylation is 2.5 and DV/K is 1.0. For -butyrobetaine hydroxylase fromPseudomonas sp. AK 1, 2-oxoglutarate decarboxylation exceeds hydroxylation with 10% when deuterated -butyrobetaine is used. No excess was found with unlabeled substrate and no internal isotope effect could be calculated. DV for the bacterial enzyme is 6. 相似文献
2.
Seedlings ofAlnus incana (nodulated and non-nodulated) andBetula papyrifera were fertilized with varying amounts (0, 10, 25, 50, 100, 250 and 500 g N g–1 soil) of labelled ammonium-N or nitrate-N ( 5.2 A% excess15N as ammonium sulphate or potassium nitrate). After 4 months in the greenhouse,15N excess in the plants were determined and an isotope dilution equation was applied to determine the percent of biomass N fixed by theA. incana/Frankia system. When ammonium was used as the sole N source and birch as the non-fixing reference, N-fixation accounted for 95%, 87% and 60% of the plant nitrogen yields with 10, 25 and 50 g N g–1 rates, additions respectively. At the 100 g N g–1 fertilization and above N-fixation accounted for less than 10% of the N yield. Similar results were obtained when non-nodulatedA. incana was used as non-fixing reference. With nitrate as the sole N source, N-fixation accounted for 98%, 97%, 97%, 86%, 56% and 12% of N yield with 10, 25, 50, 100, 250 and 500 g N g–1 additions respectively. These values were similar for both types of reference plants. The direct isotope dilution method was compared to that of the total nitrogen difference method. There was good agreement between the two methods up to 50 g N g–1 for ammonium and up to 100 g N g–1 for nitrate. The difference method produced negative values at high concentrations of nitrogen fertilization. Again similar results were obtained by the two reference plants. The results indicate that birch can be used as a non-fixing control in isotope dilution studies but that care must be exercised in selecting the type and quantity of labelled nitrogen fertilizer. 相似文献
3.
Daryl Codron Jacqui Codron Julia A. Lee-Thorp Matt Sponheimer Darryl de Ruiter James S. Brink 《European Journal of Wildlife Research》2007,53(3):161-170
This paper characterizes predator–prey interactions amongst African mammals from C4 savanna environments using stable carbon and nitrogen isotope proxies for diet. Stable carbon (δ13C) and nitrogen (δ15N) isotope data from hair and faeces of large African mammal carnivores, and herbivores as potential prey, are presented for
a diverse range of taxa. Carbon-isotope data imply that most carnivores from the “lowveld” savanna of South Africa form part
of C4 grass-based food webs. Nitrogen isotope data show clear differences between trophic levels, although it appears that the
magnitude of these differences varies between predators feeding on invertebrates and vertebrates, respectively. Whilst the
number of carnivore samples for which data are available is relatively few, and data for prey are restricted mainly to large
ungulate herbivores, results clearly demonstrate the potential for future applications of this technique to predator–prey
food webs in African savannas. In tandem with traditional approaches, stable isotopes can help elucidate patterns of predator
impacts on prey populations, domestic livestock, and resolving similar food webs in palaeoenvironmental contexts. 相似文献
4.
High δ15N of tree foliage in forests subject to high N supply has been attributed to 15N enrichment of plant available soil N pools after losses of N through processes involving N isotope fractionation (ammonia volatilization, nitrification followed by leaching and denitrification, and denitrification in itself). However, in a long-term experiment with high annual additions of NH4NO3, we found no change in the weighted average δ15N of the soil, but attributed the high δ15N of trees to loss of ectomycorrhizal fungi and their function in tree N uptake, which involves redistribution of N isotopes in the ecosystem (Högberg et al. New Phytol 189:515–525, 2011), rather than a loss of isotopically light N. Here, we compare the effects of additions of urea and NH4NO3 on the δ15N of trees and the soil profile, because we have previously found higher δ15N in tree foliage in trees in the urea plots. Doing this, we found no differences between the NH4NO3 and urea treatments in the concentration of N in the foliage, or the amounts of N in the organic mor-layer of the soil. However, the foliage of trees receiving the highest N loads in the urea treatment were more enriched in 15N than the corresponding NH4NO3 plots, and, importantly, the weighted average δ15N of the soil showed that N losses had been associated with fractionation against 15N in the urea plots. Thus, our results in combination with those of Högberg et al. (New Phytol 189:515–525, 2011) show that high δ15N of the vegetation after high N load may be caused by both an internal redistribution of the N isotopes (as a result of change of the function of ectomycorrhiza) and by losses of isotopically light N through processes fractionating against 15N (in case of urea ammonia volatilization, nitrification followed by leaching and denitrification). 相似文献
5.
《Palaeoworld》2022,31(2):249-257
The early–middle Frasnian boundary interval of the northern part of the East European Platform (northwest of Russia) corresponding to the transitans-punctata isotope event is revealed by biostratigraphically constrained conodont carbon stable isotope data (δ13Ccon). The dynamics of δ13Ccon demonstrate a three-fold pattern with positive peaks at the onset of the main phase of the transitans-punctata isotope event (upper part of Montagne Noire 4 conodont Zone, MN4; up to -22.5‰ VPDB), and during the late part of the event (lower and middle parts of MN6 Zone; up to -24.0‰ VPDB and -22.1‰ VPDB). The stratigraphic level near the MN5 and MN6 boundary is marked by a prominent negative excursion in δ13Ccon (down to -31.8‰ VPDB) that resembles the negative excursion in the terminal phase of the transitans-punctata isotope event worldwide. The δ13Ccon variations in different taxa are generally consistent but demonstrate some differences in values and amplitudes. It is assumed that variations in the carbon isotopic compositions of conodonts were mainly controlled by changes in the isotope composition of the planktons as the main food source for conodonts. 相似文献
6.
Matt K. Broadhurst Camila Domit Tailisi H. Trevizani Vincent Raoult Russell B. Millar 《Journal of fish biology》2019,95(2):589-593
We determined stable-isotope ratios for replicate muscle tissues in 13 gravid Mobula kuhlii cf. eregoodootenkee (110.4–120.4 cm disc width; WD) and their embryos (7.0–42.3 cm WD) and also yolks and histrotroph, to assess the potential implications for juvenile nutrition and habitat use. Irrespective of their development in the uterus, embryos had similar δ13C values in their muscle tissue as the mothers and both had greater values than in the histotroph. During gestation, δ13C values increased across all sample types. However, while embryo muscle tissue and the histotroph were associated with increasing 15N levels during embryonic development, this was depleted in the mothers’ muscle tissue and yolk. Although speculative, the observed variation in stable-isotope ratios might imply a dietary shift among gravid females during their early gestation. Irrespective of the underlying mechanisms, the results indicate neonates will have relatively greater δ15N values than post-partum females, which would probably confound juvenile foraging-ecology estimates. 相似文献
7.
Anne H. ODonnell Xiaojie Yao Larry D. Byers 《Biochimica et Biophysica Acta - Proteins and Proteomics》2004,1703(1):63-67
The solvent kinetic isotope effects (SKIE) on the yeast α-glucosidase-catalyzed hydrolysis of p-nitrophenyl and methyl-d-glucopyranoside were measured at 25 °C. With p-nitrophenyl-d-glucopyranoside (pNPG), the dependence of kcat/Km on pH (pD) revealed an unusually large (for glycohydrolases) solvent isotope effect on the pL-independent second-order rate constant, DOD(kcat/Km), of 1.9 (±0.3). The two pKas characterizing the pH profile were increased in D2O. The shift in pKa2 of 0.6 units is typical of acids of comparable acidity (pKa=6.5), but the increase in pKa1 (=5.7) of 0.1 unit in going from H2O to D2O is unusually small. The initial velocities show substrate inhibition (Kis/Km~200) with a small solvent isotope effect on the inhibition constant [DODKis=1.1 (±0.2)]. The solvent equilibrium isotope effects on the Kis for the competitive inhibitors d-glucose and α-methyl d-glucoside are somewhat higher [DODKi=1.5 (±0.1)]. Methyl glucoside is much less reactive than pNPG, with kcat 230 times lower and kcat/Km 5×104 times lower. The solvent isotope effect on kcat for this substrate [=1.11 (±0. 02)] is lower than that for pNPG [=1.67 (±0.07)], consistent with more extensive proton transfer in the transition state for the deglucosylation step than for the glucosylation step. 相似文献
8.
9.
The detection of steroids originating from synthetic precursors in relation to their chemically identical natural analogues has proven to be a significant challenge for doping control laboratories accredited by the World Anti-Doping Agency (WADA). Endogenous steroid abuse may be confirmed by utilising the atomic specificity of gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) that enables the precise measurement of differences in stable isotope ratios that arise as a result of fractionation patterns inherent in the source of steroids. A comprehensive carbon isotope ratio (δ13C) profiling study (n = 1262) of urinary ketosteroids is reported that demonstrates the inter-individual variation that can be expected from factors such as diet, ethnicity, gender and age within and between different populations (13 countries). This δ13C distribution is shown by principal component analysis (PCA) to provide a statistical comparison to δ13C values observed following administration of testosterone enanthate. A limited collection of steroid diol data (n = 100; consisting of three countries) is also presented with comparison to δ13C values of excreted testosterone to validate criteria for WADA accredited laboratories to prove doping offences. 相似文献
10.
Yu. A. Zolotarev E. V. Bocharov A. K. Dadayan I. E. Kasheverov M. N. Zhmak I. V. Maslennikov Yu. A. Borisov A. S. Arseniev N. F. Myasoedov V. I. Tsetlin 《Russian Journal of Bioorganic Chemistry》2000,26(9):527-531
Tritium-labeled α-conotoxin G1 with a molar radioactivity of 35 Ci/mmol and full biological activity (according to the binding
to nicotinic acetylcholine receptor) was obtained by the high-temperature solid-state catalytic isotope exchange (HSCIE).
The tritium distribution in the molecule of α-conotoxin G1 was revealed by3H NMR spectroscopy. Tritium was found in all amino acid residues except for the Asn4-Pro5-Ala6 fragment. The data on the comparative
reactivity of C-H bonds, theab initio quantum-chemical calculation of the hydrogen exchange reaction, and the information on the spatial structures of α-conotoxin
G1 in solution and in crystal state allowed us to establish that the reactivity of H atoms may be increased by their interaction
with the electron donor O and N atoms at the transition state of the HSCIE reaction. A decrease in the rate of the HSCIE reaction
could be caused by both a poor spatial accessibility of C-H bonds and a limited mobility of the peptide fragment containing
these bonds. 相似文献
11.
《Free radical biology & medicine》1999,26(1-2):129-135
A previously developed method for quantitative determination of 8-hydroxyguanine by gas chromatography-mass spectrometry was modified to allow measurement of 8-hydroxy-2′-deoxyguanosine in human urine. [4,5,6,8-13C4]8-Hydroxy-2′-deoxyguanosine was prepared by enzymatic coupling of [4,5,6,8-13C4]8-hydroxyguanine to deoxyribose-1-phosphate. Samples of human urine (2 ml) were spiked with the labeled nucleoside (13 nmol) and subjected to solid phase extraction and reversed phase high performance liquid chromatography. The 8-hydroxy-2′-deoxyguanosine thus isolated was hydrolyzed by treatment with aqueous formic acid, and the resulting 8-hydroxyguanine was converted into its tetrakis-trimethylsilyl derivative and subjected to gas-liquid chromatographic-mass spectrometric analysis. Repeated determinations of 8-hydroxy-2′-deoxyguanosine in pools of urine showed coefficients of variation of 5 and 8% at concentrations of 8-hydroxy-2′-deoxyguanosine equal to 18 and 2 nM, respectively. Determination of 8-hydroxy-2′-deoxyguanosine in samples of urine spiked with different amounts of the unlabeled nucleoside showed a mean recovery of 102%. Application of the analytical method to a group of 11 apparently healthy subjects (mean age, 47 years) showed a mean level of endogenously produced 8-hydroxy-2′-deoxyguanosine equal to 1.33 ± 0.29 μmol/mol creatinine. The level recorded for another group of 15 younger subjects (mean age, 28 years) was somewhat higher, that is, 1.58 ± 0.84 μmol/mol creatinine, corresponding to a 24-h production rate of 8-hydroxy-2′-deoxyguanosine equal to 20.6 ± 11.6 nmol (288 ± 140 pmol/24 h · kg body weight). Hemochromatosis is a hereditary disease characterized by increased absorption of iron from the gastrointestinal tract and deposition of iron in organs. Application of the analytical method to a group of 12 patients with hereditary hemochromatosis who were under treatment with venesections showed a mean level of urinary 8-hydroxy-2′-deoxyguanosine equal to 1.39 ± 0.40 μmol/mol creatinine. This value was not significantly different from those of healthy subjects. The fact that these patients had only slight or moderate iron overload at the time of urinary sample collection may have influenced the urinary levels of 8-hydroxy-2′-deoxyguanosine in the present study. 相似文献
12.
13.
Stable carbon isotope ratio of tree leaves, boles and fine litter in a tropical forest in Rondônia, Brazil 总被引:2,自引:0,他引:2
L. A. Martinelli S. Almeida I. F. Brown M. Z. Moreira R. L. Victoria L. S. L. Sternberg C. A. C. Ferreira W. W. Thomas 《Oecologia》1998,114(2):170-179
Leaves of 208 trees were collected for isotopic analysis together with wood from 36 tree boles and 18 samples of fine litter from a terra-firme forest located at Samuel Ecological Reserve, Rondônia State, in the southwestern Amazon region. The range of δ13C values in leaves was from ?28 to ?36‰, with an average (±1 SD) of ?32.1?±?1.5‰, which was more negative than the δ13C values of bole samples (?28.4?±?2.0‰) and fine litter (?28.7?±?2.0‰). These values are within the range found for tropical and subtropical forests. Pooling the δ13C values for leaf samples from trees of the same height gave averages which were positively correlated with plant height at a highly significant level, with a slope of 0.06 and an intercept of ?33.3‰ and a correlation coefficient r 2=0.70 (P<0.001). 相似文献
14.
《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》1997,692(1):7-14
Liquid chromatography–thermospray mass spectrometry (LC–TSP-MS) using isotope dilution was investigated for quantitative analysis of cortisol, cortisone, prednisolone and prednisone in human plasma. Complete separation attained by a LiChroCART Supersupher reversed-phase column and elution with 0.05 M ammonium formate–tetrahydrofuran–methanol (180:53:17, v/v/v) resulted in a significantly large isotope effect of the deuterium-labeled analogs on the HPLC behavior and caused difficulty in quantification. Reduction of the isotope effect on the retention times using 0.05 M ammonium formate–acetonitrile (65:35, v/v) permitted accurate quantification of cortisol and cortisone by the isotope dilution LC–TSP-MS, although separation between cortisol and prednisone was incomplete. 相似文献
15.
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16.
Yi Wang Jiabao Xu Lingchao Kong Tang Liu Lingbo Yi Hongjuan Wang Wei E. Huang Chunmiao Zheng 《Microbial biotechnology》2020,13(2):572-583
Human intestinal microbiota is important to host health and is associated with various diseases. It is a challenge to identify the functions and metabolic activity of microorganisms at the single-cell level in gut microbial community. In this study, we applied Raman microspectroscopy and deuterium isotope probing (Raman–DIP) to quantitatively measure the metabolic activities of intestinal bacteria from two individuals and analysed lipids and phenylalanine metabolic pathways of functional microorganisms in situ. After anaerobically incubating the human faeces with heavy water (D2O), D2O with specific substrates (glucose, tyrosine, tryptophan and oleic acid) and deuterated glucose, the C–D band in single-cell Raman spectra appeared in some bacteria in faeces, due to the Raman shift from the C–H band. Such Raman shift was used to indicate the general metabolic activity and the activities in response to the specific substrates. In the two individuals' intestinal microbiota, the structures of the microbial communities were different and the general metabolic activities were 76 ± 1.0% and 30 ± 2.0%. We found that glucose, but not tyrosine, tryptophan and oleic acid, significantly stimulated metabolic activity of the intestinal bacteria. We also demonstrated that the bacteria within microbiota preferably used glucose to synthesize fatty acids in faeces environment, whilst they used glucose to synthesize phenylalanine in laboratory growth environment (e.g. LB medium). Our work provides a useful approach for investigating the metabolic activity in situ and revealing different pathways of human intestinal microbiota at the single-cell level. 相似文献
17.
Xinhua Dai Xiang FangFuhai Su Mengrui YangHongmei Li Jian ZhouRuifeng Xu 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2010,878(19):1634-1638
A high order method for measuring urea concentrations in milk and milk powder was developed. The method can be applied to certify the concentration of urea in some new milk and milk powder CRMs. This high accurate method for analysis of milk is valuable given the inherent challenges associated with the complexity of the sample matrix. A measurement procedure based on gas chromatography/isotope dilution mass spectrometry (GC/IDMS) was developed. Samples were pre-treated with acetonitrile to remove proteins and the method was applied to determine urea concentrations in milk and milk powder. Excellent precision was obtained, with within- and between-set coefficients of variation of 0.15–0.46 and 0.18–0.65%, respectively. The measurement uncertainty is evaluated. The method can trace to mass. 相似文献
18.
Variation in the oxygen isotope ratio of phloem sap sucrose from castor bean. Evidence in support of the Péclet effect 总被引:1,自引:0,他引:1
Theory suggests that the level of enrichment of (18)O above source water in plant organic material (Delta) may provide an integrative indicator of control of water loss. However, there are still gaps in our understanding of the processes affecting Delta. One such gap is the observed discrepancy between modeled enrichment of water at the sites of evaporation within the leaf and measured enrichment of the leaf water as a whole (Delta(L)). Farquhar and Lloyd (1993) suggested that this may be caused by a Péclet effect. It is also unclear whether organic material formed in the leaf reflects enrichment of water at the sites of evaporation within the leaf or Delta(L). To investigate this question castor bean (Ricinus communis L.) leaves, still attached to the plant, were sealed into a controlled-environment gas exchange chamber and subjected to a step change in leaf-to-air vapor pressure difference. Sucrose was collected from a cut on the petiole of the leaf in the chamber under equilibrium conditions and every hour for 6 h after the change in leaf-to-air vapor pressure difference. Oxygen isotope composition of sucrose in the phloem sap (Delta(suc)) reflected modeled Delta(L). A model is presented describing Delta(suc) at isotopic steady state, and accounts for 96% of variation in measured Delta(suc). The data strongly support the Péclet effect theory. 相似文献
19.
Craig Polson Joseph L. Cantone Cong Wei Dieter M. Drexler Jere E. Meredith Jr. 《Analytical biochemistry》2014
Microtubules (MTs) are highly dynamic polymers composed of α- and β-tubulin heterodimers. Dysregulation of MT dynamics in neurons may be a contributing factor in the progression of various neurodegenerative diseases. We developed a stable isotope labeling by amino acids in cell culture (SILAC)-based liquid chromatography–mass spectrometry (LC–MS) method to measure the fraction of [13C6]leucine-labeled α-tubulin-derived surrogate peptides. Using this approach, we measured the time course of incorporation of [13C6]leucine label into the MT and dimer pools isolated from cycling cells and rat primary hippocampal neurons. We found that the MT pool is in rapid equilibrium with the dimer pool in the cycling cells, consistent with rapid MT polymerization/depolymerization during cell proliferation. Conversely, in neurons, we found that labeling of the MT pool was rapid, whereas the dimer pool was delayed. These results suggest that newly synthesized α-tubulin is first incorporated into MTs or complexes that co-sediment with MTs and that appearance of labeled α-tubulin in the dimer pool may be a consequence of MT depolymerization or breakdown. Our results demonstrate that a SILAC-based approach can be used to measure MT dynamics and may have utility for exploring MT dysregulation in various models of neurodegenerative disease. 相似文献
20.
Philipp Hoffmann Michaela F. Hartmann Thomas Remer Klaus-Peter Zimmer Stefan A. Wudy 《Steroids》2010,75(13-14):1067-1074
Oestrogens, such as oestrone (E1), 17β-oestradiol (E2), oestriol (E3) and their biologically active metabolites 2-methoxyoestrone (2-MeOE1), 2-hydroxyoestradiol (2-OHE2) 16-ketooestradiol (16-OE2), 16-epioestriol (16-epiE3), as well as testosterone (T) play an important role in physiological and pathological developmental processes during human development. We therefore aimed at developing an isotope dilution/bench top gas chromatography–mass spectrometry (ID/GC–MS) method, based on benchtop GC–MS, for the simultaneous determination (‘profiling’) of the above analytes in children. The method consisted of equilibration of urine (5 ml) with a cocktail containing stable isotope-labelled analogues of the analytes as internal standards ([2,4-2H2]E1, [2,4,16,16-2H4]E2, [2,4,17-2H3]E3, [16,16,17-2H3]T, [1,4,16,16-2H4]2-MeOE1, [1,4,16,16,17-2H5]2-OHE2, [2,4,15,15,17-2H5]16-OE2 and [2,4-2H2]16-epiE3). Then, solid-phase extraction (C18 cartridges), enzymatic hydrolysis (sulphatase from Helix pomatia (type H-1)), re-extraction, purification by anion exchange chromatography and derivatisation to trimethylsilyl ethers followed. The samples were analysed by GC–MS (Agilent GC 6890N/5975MSD; fused silica capillary column 25 m × 0.2 mm i.d., film 0.10 μm). Calibration plots were linear and showed excellent reproducibility with coefficients of determination (r2) between 0.999 and 1.000. Intra- and inter-assay coefficients of variation (CV) were <2.21% for all quantified metabolites. Sensitivity was highest for 2-OHE2 (0.25 pg per absolute injection: signal-to-noise ratio (S/N) = 3) and lowest for 16-epiE3 (2 pg per absolute injection: S/N = 2.6), translating into corresponding urine sample analyte concentrations of 0.025 ng ml?1 and 0.2 ng ml?1, respectively. Accuracy – determined in a two-level spike experiment – showed relative errors ranging between 0.15% for 16-OE2 and 11.63% for 2-OHE2. Chromatography showed clear peak shapes for the components analysed. In summary, we describe a practical, sensitive and specific ID/GC–MS assay capable of profiling the above-mentioned steroids in human urine from childhood onwards. 相似文献