Large old trees influence patterns of delta13C and delta15N in forests

Large old trees are the dominant primary producers of native pine forest, but their influence on spatial patterns of soil properties and potential feedback to tree regeneration in their neighbourhood is poorly understood. We measured stable isotopes of carbon (delta(13)C) and nitrogen (delta(15)N) in soil and litter taken from three zones of influence (inner, middle and outer zone) around the trunk of freestanding old Scots pine (Pinus sylvestris L.) trees, to determine the trees' influence on below-ground properties. We also measured delta(15)N and delta(13)C in wood cores extracted from the old trees and from regenerating trees growing within their three zones of influence. We found a significant and positive gradient in soil delta(15)N from the inner zone, nearest to the tree centre, to the outer zone beyond the tree crown. This was probably caused by the higher input of (15)N-depleted litter below the tree crown. In contrast, the soil delta(13)C did not change along the gradient of tree influence. Distance-related trends, although weak, were visible in the wood delta(15)N and delta(13)C of regenerating trees. Moreover, the wood delta(15)N of small trees showed a weak negative relationship with soil N content in the relevant zone of influence. Our results indicate that large old trees control below-ground conditions in their immediate surroundings, and that stable isotopes might act as markers for the spatial and temporal extent of these below-ground effects.     

Increasing the sensitivity of delta13C and delta15N abundance measurements by a high sensitivity elemental analyzer connected to an isotope ratio mass spectrometer

A common elemental analyzer system connected to a temperature-controlled gas chromatography (GC) column and coupled to an isotope ratio mass spectrometer was improved to decrease the determination limit for a simultaneous stable isotope ratio measurement of nitrogen and carbon dioxide. The additional use of a special ashtray system to collect the combustion residuals permitted more time-efficient work. These modifications to the elemental analyzer allowed precise measurements to be made down to 1.5 microg nitrogen and 10 microg carbon for stable isotope analysis. Low system background values and an acceptable signal-to-noise ratio have made an additional blank correction for these low sample measurements unnecessary. We provide a precision of this stable isotope analysis for lowest amounts of 1.2-2 microg nitrogen with a standard deviation of +/-0.496 per thousand (n = 27) and for 8.2-15 microg carbon with a standard deviation of +/-0.257 per thousand (n = 31) across different sample runs under stipulated conditions. This application can be established in an automatic mode without cryofocusing procedures.     

Isotope ratio mass spectrometry: delta13C and delta15 N analysis for tracing the origin of illicit drugs

Gas chromatography/combustion/mass spectrometry (GC-C-MS) and elemental analysis/mass spectrometry (EA-MS) techniques are proposed to estimate delta(13)C and delta(15)N values in heroin, morphine, cocaine and hemp leaves, for the purposes of tracing the geographical origins of seized drugs. The values of isotope ratios for pure drugs and drugs with impurities were compared. It was demonstrated that large samples (up to 3 x 10(-6) g C) were combusted completely, so that the results obtained were valid. The data are considered to be an essential supplement to a wide-scale database designed specifically for the delta(13)C and delta(15)N values of drugs. The potential forensic and academic significance of the results is discussed.     

Tissue and fixative dependent shifts of delta13C and delta15N in preserved ecological material

Carbon and nitrogen stable isotope analyses are routinely used to investigate aquatic food webs, and have potential application in retrospective investigations using archived materials. However, such analyses assume that storage does not alter isotopic signatures of materials preserved, or that changes in isotopic composition during storage are predictable. Here we examine preservation shifts on cod (Gadus morhua) muscle, roe and liver tissue over 21 months following preservation in 80% ethanol, in 4% formaldehyde, and by freezing. Preservation shifts were not consistent among tissues. High protein tissues exhibited greater delta(15)N shifts than low protein tissues in 4% formaldehyde, while greater delta(13)C shifts occurred in relatively higher fat tissues when preserved in alcohol. Freezing did not change isotopic signatures. Responses of delta(15)N and delta(13)C are explained by differences in the preservative's isotopic signature and the reaction properties and biochemical composition of the tissues preserved. The results clarify some of the processes that lead to isotopic change during preservation.     

Two new marine sediment standard reference materials (SRMs) for the determination of organic contaminants

Two new marine sediment standard reference materials (SRMs), SRM 1941b Organics in Marine Sediment and SRM 1944 New York/New Jersey Waterway Sediment, have been recently issued by the National Institute of Standards and Technology (NIST) for the determination of organic contaminants including polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyl (PCB) congeners, and chlorinated pesticides. Both sediment SRMs were analyzed using multiple analytical methods including gas chromatography/mass spectrometry (GC/MS) on columns with different selectivity, reversed-phase liquid chromatography with fluorescence detection (for PAHs only), and GC with electron capture detection (for PCBs and pesticides only). SRM 1941b has certified concentrations for 24 PAHs, 29 PCB congeners, and 7 pesticides, and SRM 1944 has certified concentrations for 24 PAHs, 29 PCB congeners, and 4 pesticides. Reference concentrations are also provided for an additional 58 (SRM 1941b) and 39 (SRM 1944) PAHs, PCB congeners, and pesticides. SRM 1944, which was collected from multiple sites within New York/New Jersey coastal waterways, has contaminant concentrations that are generally a factor of 10–20 greater than SRM 1941b, which was collected in the Baltimore (Maryland) harbor. These two SRMs represent the most extensively characterized marine sediment certified reference materials available for the determination of organic contaminants.Electronic Supplementary Material Supplementary material is available in the online version of this article at . A link in the frame on the left on that page takes you directly to the supplementary material.     

Methods to adjust for the interference of N2O on delta13C and delta18O measurements of CO2 from soil mineralization

In this paper we present an overview of the present knowledge relating to methods that avoid interference of N2O on delta13C and delta18O measurements of CO2. The main focus of research to date has been on atmospheric samples. However, N2O is predominantly generated by soil processes. Isotope analyses related to soil trace gas emissions are often performed with continuous flow isotope ratio mass spectrometers, which do not necessarily have the high precision needed for atmospheric research. However, it was shown by using laboratory and field samples that a correction to obtain reliable delta13C and delta18O values is also required for a commercial continuous flow isotope ratio mass spectrometer. The capillary gas chromatography column of the original equipment was changed to a packed Porapak Q column. This adaptation resulted in an improved accuracy and precision of delta13C (standard deviation(Ghent): from 0.2 to 0.08 per thousand; standard deviation(Lincoln): from 0.2 to 0.13 per thousand) of CO2 for N2O/CO2 ratios up to 0.1. For delta18O there was an improvement for the standard deviation measured at Ghent University (0.13 to 0.08 per thousand) but not for the measurements at Lincoln University (0.08 to 0.23 per thousand). The benefits of using the packed Porapak Q column compared with the theoretical correction method meant that samples were not limited to small N(2)O concentrations, they did not require an extra N2O concentration measurement, and measurements were independent of the variable isotopic composition of N2O from soil.     

1H, 13C and 15N NMR spectral assignments for new triazapentalene derivatives

Mesomeric heteropentalene betaines are conjugated fused polyheterocyclic structures that represent interesting intermediates for organic synthesis. Five such structures, containing at least four nitrogen atoms and various substituents, have been characterized by ¹H, ¹³C and ¹⁵N NMR. We report, apparently for the first time, nitrogen NMR data and coupling information on such systems. Inter‐ring long‐range correlations across five bonds with ¹⁵N (⁵J_HN) and up to seven bonds with ¹³C (⁶J_HC and ⁷J_HC) were observed in HSQC experiments. The incorporation of an electron‐withdrawing substituent such as NO₂ was observed to cause an increase in the magnitude of the remote couplings and deshielding of nearby protons, carbons and on all nitrogen atoms of the structure, including remote ones situated on other cycles. Copyright © 2009 John Wiley & Sons, Ltd.     

A comparison of muscle- and scale-derived delta13C and delta15N across three life-history stages of Atlantic salmon, Salmo salar

Stable isotope signatures were obtained from paired scale and muscle tissue samples from smolt, post-smolt and one-sea-winter adult Atlantic salmon (Salmo salar). Post-smolt and adult scales were separated into central and outer (marine) portions with analyses carried out on the marine growth section of both life-history stages and the central portion for the adult scales. Muscle and scale delta(13)C and delta(15)N signatures were assessed (1) to determine whether a linear relationship exists between tissue types, (2) to determine if a constant offset exists between tissue signatures across all life-history stages, and (3) to evaluate whether underplating imparts a significant bias to life-history scale segments that would preclude their use in retrospective analyses of any ontogenetic dietary changes between life-history stages. Significant correlations were found to exist between muscle and scale stable isotope signatures obtained from smolts (delta(13)C and delta(15)N) and adults (delta(15)N). Both the muscle and the scale signatures captured the dietary shift associated with the transition from freshwater to the marine environment. Post-smolt and adult scales were depleted relative to muscle tissue, which may be attributed to isotopic differences in amino acid composition between muscle and scale tissues. The results suggest that scales may better represent dietary carbon sources because they are not influenced by lipid dynamics. The scale, however, appears less responsive to short-term shifts in diet relative to muscle and, therefore, must be used only to infer seasonally integrated dietary patterns for slow-growing life-history stages. Copyright (c) 2008 John Wiley & Sons, Ltd.     

Applying the silver‐tube introduction method for thermal conversion elemental analyses and a new δ2H value for NBS 22 oil

The δ²H_VSMOW–SLAP value of total hydrogen of the international measurement standard NBS 22 oil was determined by a new method of sealing water in silver tubes for use in a thermal conversion elemental analysis (TC/EA) reduction unit. The isotopic fractionation of water due to evaporation is virtually non‐existent in this silver‐tube method. A new value for the δ²H_VSMOW–SLAP of NBS 22 oil, calibrated with isotopic reference waters, was determined to be ?116.9 ± 0.8‰ (1σ and n = 31). Published in 2010 by John Wiley & Sons, Ltd.     

Clinical-scale investigation of stable isotopes in human blood: delta13C and delta15N from 406 patients at the Johns Hopkins Medical Institutions

Objective chemical biomarkers are needed in clinical studies of diet-related diseases to supplement subjective self-reporting methods. We report on several critical experiments for the development of clinically legitimate dietary stable isotope biomarkers within human blood. Our examination of human blood revealed the following: (1) Within blood clot and serum from anonymous individuals (201 males, 205 females) we observed: mean serum delta13C = -19.1 +/- 0.8 per thousand (standard deviation, SD); clot, -19.3 +/- 0.8 per thousand (SD); range = -15.8 per thousand to -23.4 per thousand. Highly statistically significant differences are observed between clot and serum, males and females for both clot and serum. For 15N (n = 206), mean serum = +8.8 +/- 0.5 per thousand (SD); clot +7.4 +/- 0.4 per thousand (SD); range = +6.3 per thousand to +10.5 per thousand. Blood serum is enriched in 15N relative to blood clot by +1.4 per thousand on average, which may reflect differing protein amino acid content. Serum nitrogen is statistically significantly different for males and females, however, clot shows no statistical difference. (2) Relative to clot, capillary blood is marginally different for 13C, but not 15N. Clot 13C is not significantly different from serum; however, it is depleted in 15N by 1.5 per thousand relative to serum. (3) We assessed the effect of blood additives (sodium fluoride and polymerized acrylamide resin) and laboratory process (autoclaving, freeze drying) commonly used to preserve or prepare venous blood. On average, no alteration in delta13C or delta15N is detected compared with unadulterated blood from the same individual. (4) Storage of blood with and without the additives described above for a period of up to 115 days exhibits statistically significant differences for 13C and 15N for sodium fluoride. However, storage for unadulterated blood and blood preserved with polymerized acrylamide resin does not change the delta13C or delta15N isotopic composition of the blood in a significant way. With these experiments, we gain a clinical context for future development of a stable isotope based dietary biomarker.     

Co-acquisition of hyperpolarised 13C and 15N NMR spectra

Recent developments in dynamic nuclear polarisation now allow significant enhancements to be generated in the cryo solid state and transferred to the liquid state for detection at high resolution. We demonstrate that the Ardenkjaer-Larsen method can be extended by taking advantage of the properties of the trityl radicals used. It is possible to hyperpolarise 13C and 15N simultaneously in the solid state, and to maintain these hyperpolarisations through rapid dissolution into the liquid state. We demonstrate the almost simultaneous measurement of hyperpolarised 13C and hyperpolarised 15N NMR spectra. The prospects for further improvement of the method using contemporary technology are also discussed.     

Diode laser spectroscopy of the fundamental bands of 12C14N, 13C14N, 12C15N, 13C15N free radicals in the ground 2 Sigma+ electronic state

Rotationally resolved spectra of the fundamental band of the CN free radical in four isotopic forms have been measured using tunable diode laser absorption spectroscopy. The source of the radical was a microwave discharge in a mixture of isotopically selected methane and nitrogen diluted with argon. The lines were measured to an accuracy of 5 x 10(-4) cm(-1) and fitted to the formula for the vibration rotation spectrum of a diatomic molecule, including quartic distortion constants. The band origins of each of the isotopomers from the five parameter fits were found to be 12C14N: 2042.42115(38) cm(-1), 13C14N: 2000.08479(23) cm(-1), 12C15N: 2011.25594(25) cm(-1), 13C15N: 1968.22093(33) cm(-1) with one standard deviation from the fit given in parenthesis. Some of the lines showed a resolved splitting due to the spin rotation interaction. This was averaged for fitting purposes. The average equilibrium internuclear distance derived from the upsilon = 0 and 1 rotational constants of the four isotopomers is 1.171800(6) A which is in good agreement with the value determined from microwave spectroscopy.     

Chemical shielding anisotropy of 13C and 15N in acetonitrile

High resolution ¹³C and ¹⁵N NMR spectra have been obtained for powdered CH₃CN. The presence of resolved dipolar structure in the ¹³C spectra permits the conclusion that the symmetry axis of the ¹³C shielding tensor lies along the CN bond direction.     

13C and 15N NMR chemical shifts of alkylsubstituted benzonitriles

¹³C NMR data of 14 and ¹⁵N NMR data of four alkylsubstituted benzonitriles are reported and discussed in relation to substituent effects and stereochemistry.     

Characterisation of uniformly 13C, 15N labelled bacteriochlorophyll a and bacteriopheophytin a in solution and in solid state: complete assignment of the 13C, 1H and 15N chemical shifts

In this investigation we report a complete assignment of (13)C, (1)H and (15)N solution and solid state chemical shifts of two bacterial photosynthetic pigments, bacteriochlorophyll (BChl) a and bacteriopheophytin (BPheo) a. Uniform stable-isotope labelling strategies were developed and applied to biosynthetic preparation of photosynthetic pigments, namely uniformly (13)C, (15)N labelled BChl a and BPheo a. Uniform stable-isotope labelling with (13)C, (15)N allowed performing the assignment of the (13)C, (15)N and (1)H resonances. The photosynthetic pigments were isolated from the biomass of photosynthetic bacteria Rhodopseudomonas palustris 17001 grown in uniformly (13)C (99%) and (15)N (98%) enriched medium. Both pigments were characterised by NMR in solution (acetone-d(6)) and by MAS NMR in solid state and their NMR resonances were recorded and assigned through standard liquid 2D (13)C-(13)C COSY, (1)H-(13)C HMQC, (1)H-(15)N HMBC and solid 2D (13)C-(13)C RFDR, (1)H-(13)C FSLG HETCOR and (1)H-(15)N HETCOR correlation techniques at 600 MHz and 750 MHz. The characterisation of pigments is of interest from biochemical to pharmaceutical industries, photosynthesis and food research.     

Site-specific 13C chemical shift anisotropy measurements in a uniformly 15N,13C-labeled microcrystalline protein by 3D magic-angle spinning NMR spectroscopy

In this Communication, we introduce a 3D magic-angle spinning recoupling experiment that correlates chemical shift anisotropy (CSA) powder line shapes with two dimensions of site-resolved isotropic chemical shifts. The principal tensor elements from 127 ROCSA line shapes are reported, constraining 102 unique backbone and side-chain 13C sites in a microcrystalline protein (the 56 residue beta1 immunoglobulin binding domain of protein G). The tensor elements, determined by fitting to numerical simulations, agree well with quantum chemical predictions. The experiments, therefore, validate calculations of CSAs in a protein of known structure. The data will be useful for the development of side-chain CSA quantum calculations and will aid in the design and interpretation of solution NMR experiments that utilize CSA-dipole cross-correlation to constrain torsion angles or to enhance resolution and sensitivity (such as in TROSY). Furthermore, the methodology described here will enable databases of CSA data to be generated with higher efficiency, for purposes of direct protein structure refinement.     

Synthesis and 13C, 15N NMR study of a new functionalized pyrido[2,3-b]indole derivative

New functionalized α-carbolinones especially the 4-hydroxy-3-nitro-1H,9H-pyrido[2,3-b]indol-2-one were synthesized in a good yield, three-step reaction. A complete ¹³C, ¹⁵N nmr study of this carbolinone and precursors is presented.