14N Ratios Research Articles

Precision and sensitivity of the measurement of 15N enrichment in D-alanine from bacterial cell walls using positive/negative ion mass spectrometry

Sensitive detection of cellular components from specific groups of microbes can be utilized as ‘signatures’ in the examination of microbial consortia from soils, sediments or biofilms. Utilizing capillary gas chromatography/mass spectrometry and stereospecific derivatizing agents, D-alanine, a component localized in the prokaryotic (bacteria) cell wall, can be detected reproducibly. Enrichments of D- 15N]alanine determined in E. coli grown with [ 15N]ammonia can be determined with precision at 1.0 atom%. Chemical ionization with methane gas and the detection of negative ions (M–HF) −· and (M−F or M+H − HF) − formed from the heptafluorobutyryl D-2 butanol ester of D-alanine allowed as little as 8 pg (90 fmol) to be detected reproducibly. This method can be utilized to define the metabolic activity in terms of 15N incorporation at the level of 10 3−10 4 cells, as a function of the 15N− 14N ratio.

Variations in the natural abundance of 15N in estuarine suspended particulate matter: A specific indicator of biological processing

In this study, the 14N: 15N ratio of suspended particulate material collected from the Tamar river estuary, south-west England, is described. Three populations of particles, distinguishable by their 15N content, were observed. This investigation has shown that populations of estuarine particles are generated by biological transformations in situ and that the 15N content of estuarine particles does not merely reflect hydrodynamic mixing of the freshwater and seawater source particulate material.

Natural Abundance of Nitrogen-15 as a Tool for Tracing Alder-Fixed Nitrogen

Ratios of 15N to 14N often differ between pools within ecosystems, and measurement of these natural-abundance ratios might allow transfers among pools to be traced. We tested this approach for its ability to trace biologically-fixed N in conifer plantations. Ratios of 15N to 14N were measured in soil total-N, ammonium, and nitrate, and in foliage of Douglas-fir [Pseudotsuga menziesii (Mirb.) Franco] and red alder (Alnus rubra Bong.) at four sites. Two ecosystems were sampled at each site, one a pure conifer stand and the other a mixed alder-conifer stand. Isotope ratios differed significantly among stands, but no pattern was consistent across all locations. Soil NH+4 at all sites (and soil nitrate at one site) was significantly depleted in 15N relative to other N pools. Isotope discrimination clearly occurs during N transfers at these sites, but the 15N natural-abundance method does not provide a simple picture of N cycling at the ecosystem level.

Stable nitrogen isotopes in zooplankton: Some geographic and temporal variations in the North Pacific1

Copepods and chaetognaths from the Southern California Bight had higher 15N:14N ratios than did those from the North Pacific Central Gyre. Within the bight, zooplankters were more enriched in 15N when NO3− was the major nitrogenous nutrient for phytoplankton than at other times. The 15N enrichment is related to the degree of exchange of nitrogen between the euphotic zone and deeper water, and the low 15N:14N ratio in the central gyre is probably also related to the importance of nitrogen fixation. In contrast, little geographic or temporal variation was observed in zooplanktonic δ13C.

Nitrogen isotopes in lunar highlands breccias

Some breccias from the lunar highlands have probably trapped solar wind gases at a very early epoch in the history of the moon, as implied by their high contents of parentless fissiogenic xenon and sometimes, of parentless radiogenic 129Xe. Four samples of this type, on which noble gas data already exist, have been selected for analysis of nitrogen contents and isotopic composition, by using step-wise heating techniques: 14047, 14055, 14307, 60255. Since uncertainties in the evolution of the solar wind 15N/ 14N ratio with time are due in part to uncertainties in the measurement of the epoch of exposure, those samples provided the opportunity to measure the isotopic composition of nitrogen which has been trapped in the remote past, avoiding the problems inherent in the use of spallogenic nuclides. Results show that, in the samples studied from the Apollo 14 landing site, nitrogen is not particularly light, and has not been acquired, as a whole, in very ancient times. The conflicting presence of both parentless xenon and nitrogen of relatively “recent” isotopic signature can be explained if the hypothetical light nitrogen is diluted by more abundant, heavier nitrogen. Accordingly, the very ancient soil components which are implied in these objects by the presence of excess fission xenon have been re-exposed at a much later epoch, or mixed with some younger soil components, before the compaction event. The present data do not question the hypothesis of a secular isotopic variation of lunar trapped nitrogen, but cannot prove that very light nitrogen was trapped together with parentless fission xenon in the soil components of the highlands soil breccias. The very unusual release pattern of nitrogen in breccia 60255 can result from nitrogen isotopic homogenization with gas loss.

Simultaneous measurements and determinations of stable carbon and nitrogen isotope ratios, and organic carbon and nitrogen contents in biological samples by coupling of a small quadrupole mass spectrometer and modified carbon-nitrogen elemental analyzer

A simple and rapid method for simultaneous measurements of 13C/ 12C and 15N/ 14N ratios and organic C and N contents in biological samples at microgram levels by coupling of QMS and modified CHN elemental analyzer was examined for the use of stable carbon and nitrogen isotope as a tracer. Monitoring of CO 2(m/z=44 and 45) and N 2(m/z=28 and 29) ion currents due to the combustion product gas by the elemental analyzer allows simultaneous measurements of both isotope ratios within 0.5 % at 10–500 microgram. Organic C and N contents were determined within 5–12 % relative standard deviation at the same levels, depending upon the combustion conditions.

The determination of 14N: 15N isotope ratios by using the (3,0) and (4,1) bandheads of the N 2 second positive system

A good linear calibration for the isotopic composition of ammonia can be obtained over the range 5–30 atom-% nitrogen-15, by using an electrodeless discharge tube excited at 2450 MHz and cooled to suppress OH emission. Reproducible relative intensities of comparable bandheads from the two transitions indicate a stable excitation temperature in the discharge tube. It is concluded that the transitions in general use, the (2,0) or (1,0), are preferable for the determination of 14N: 15N ratios.

Ages and composition of gas trapped in Allan Hills and Byrd core ice

Gas is extracted from large (6–31 kg) Antarctic ice samples to obtain sufficient CO 2 for 14C measurements with small low-level proportional counters. The 14C ages of Byrd core ice are in accord with glaciological estimates ranging from (2.2 −1.1 +1.4)×10 3 yr at 271 m depth to more than 8 × 10 3 yr at 1071 m depth. The CO 2 abundances in gas extracted from Byrd core ice range from 0.0216 to 0.051%, with below present-day atmosphere CO 2 abundances for ice from 1068 and 1469 m depths. The CO 2 abundance in gas from Allan Hills surface ice samples ranges between four and six times the atmospheric value and the CO 2 had a specific activity three times that of contemporary carbon. A possible explanation for the anomalously high specific activity is surface melting with the incorporation into CO 2 of 14C produced by cosmic ray spallation of oxygen in ice. The CO 2 abundance in gas extracted from subsurface Allan Hills ice ranged from 0.030 to 0.065%, and the specific activities are below contemporary carbon, indicating ages greater than ∼5×10 3 yr. The 18O/ 16O ratio of oxygen in the trapped gas is the same as that of atmospheric oxygen and differs markedly from the 18O/ 16O ratio in the ice. The O 2, N 2, and Ar abundances and isotopic compositions are similar to those in contemporary air, except for positive 15N/ 14N ratios in a few samples.

Nitrogen isotopes in the solar system

Measurements of the isotopic composition of nitrogen in the solar system are summarized. We show that the 30% change, during the last 3 to 4 billion years, of 15N 14N in solar-wind-bearing lunar soils and breccias probably does not reflect changes in this ratio at the solar surface. Such changes, whether by spallation or thermonuclear reactions are ruled out by comparing the yields of 15N with those of other rare isotopes such as 9Be, 11B, 3He or 13C, even if an arbitrary degree of solar mixing is introduced. Moreover, we calculate that the solar activity required for producing significant amounts of 15N by spallation at the solar surface should have resulted in a particle bombardment of the Moon of an intensity that would have produced amounts of spallation isotopes ( e.g. 15N, 21Ne, 38Ar, 131Xe) several orders of magnitude in excess of what is actually found in the whole regolith. We argue that accretion of interstellar matter also does not work as a cause for a significant change of the photospheric 15N 14N ratio. Evidence is presented that the mixing depth at the solar surface on a time scale of ≳10 9 years is (10 −2 −10 −1) M ⊙ Mixing to this depth renders accretion of interstellar matter as a source of compositional changes at the solar surface inefficient, even if allowance is made for the expected large difference in the accretion rates of condensed and gaseous matter. A quantitative treatment of several alternatives of solar accretion leads to serious contradictions ( e.g. with the low Ne abundances in planetary atmospheres or with the amounts of nitrogen that should have been directly accreted by the Moon), and we conclude that accretion during the main sequence life of the Sun is an unlikely source of changes in 15N 14N at the solar surface. A ratio of 15N 14N = (4.0 ± 0.3) × 10 −3 is our best estimate for average solar system material and for the Sun. We propose that a rare, very light nitrogen component (called LPN) is admixed in varying amounts to planetary matter. Undiluted LPN has not been found in meteorites or planetary atmospheres, but we show that the combined effects of LPN admixture and isotope fractionation can in principle account for the variability of 15N 14N observed in the planetary system. Determination of the Jovian 15N 14N ratio with an accuracy of ~10% would crucially test our interpretation of the nitrogen isotope observations.

Robinson-type nuclear quadrupole resonance spectrometer adapted to field-effect transistors

A nuclear quadrupole resonance spectrometer, the Robinson circuit adapted with field-effect transistors, has operated satisfactorily up to 30 MHz with rf levels ranging from 40 mV to 2.6 V peak to peak across the sample coil. The S/N ratio of 14N in hexamethylenetetramin is 85 at room temperature and 150 at 77 K, with a sample volume of 6 cm3 and a time constant of 10 s. The spectrometer is also capable of measuring the spin-lattice relaxation time by the saturation method.

Ancient solar wind in lunar microbreccias

The Apollo 11 soil breccias are samplers of the ancient lunar environment due to their history in the regolith and their efficient closure to addition of recent solar wind upon compaction. These breccias contain the lowest 15N/ 14N isotopic ratio yet reported for any lunar sample (in fact, for any natural sample). This extends the range of variation of 15N/ 14N of the solar wind to greater than 30%, from a δ 15N of −190‰ in the past to +120‰ at present. No mechanism is yet known that is capable of accounting for such a large change in the 15N/ 14N ratio without producing a substantial concomitant change in the 13C/ 12C ratio, although some sort of nuclear reaction in the sun appears to be required. Apollo 11 soil breccias and 15086 are all formed by meteoritic impacts which compact the lower regolith against the basement rock without much heating. Rock 15086 formed from the layer of regolith between 100 and 200 cm depth, as shown by the close agreement between the nitrogen content and isotopic ratios of 15086 and those of the Apollo 15 deep drill core. Cosmic ray exposure ages, based on spallation-produced 15N, are 2.3 ± 0.4 b.y. for Apollo 11 breccias. This age is much greater than the estimate from cosmogenic 21Ne, presumably due to diffusive loss of neon.

Nitrogen isotope fractionations in the Fischer-Tropsch synthesis and in the Miller-Urey reaction

Nitrogen isotope fractionations have been measured in Fischer-Tropsch and Miller-Urey reactions in order to determine whether these processes can account for the large 15N/ 14N ratios found in organic matter in carbonaceous chondrites. Polymeric material formed in the Fischer-Tropsch reaction was enriched in 15N by only 3‰ relative to the starting material (NH 3). The 15N enrichment in polymers from the Miller-Urey reaction was 10–12‰. Both of these fractionations are small compared to the 80–90‰ differences observed between enstatite chondrites and carbonaceous chondrites. These large differences are apparently due to temporal or spatial variations in the isotopic composition of nitrogen in the solar nebula, rather than to fractionation during the production of organic compounds.

Fractionation of nitrogen isotopes by animals: a further complication to the use of variations in the natural abundance of 15N for tracer studies

SUMMARYA study of the fractionation of nitrogen isotopes in the diet by cattle is described and the results discussed.Compared with the diet, urine had a lower ratio of 15N to 14N, but faeces, blood and milk all had a higher ratio.It is argued that the use of natural 15N as a tracer in grazed ecosystems is more complicated than was at first thought.

A simple procedure for 15N determinations in relatively large samples by emission spectrometry

Abstract Discharge tubes containing argon, and NH3 separated from atmospheric N2 by condensation in a liquid‐nitrogen—cooled trap, are used to determine 15N: 14N ratios in ammonium solutions. Reduction of NH3 to N2 during sample preparation is not required; the dissociation of NH3 in the discharge tube produces sufficiently intense N2 band spectra for the isotope analysis. The method can be readily applied to the analysis of ammonium solutions derived from Kjeldahl digests of plant material.

Influence of partial pressures of acetylene and nitrogen upon nitrogenase activity of species of Beijerinckia.

Acetylene reduction and nitrogen fixation by strains of Beijerinckia indica and B. lacticogenes increased with increased partial pressures of acetylene and nitrogen up to 80 kPa. The optical emission spectrophotometric method was used for the determination of 14N:15N ratios. The molar ratios of acetylene to nitrogen varied greatly from the theoretical value.

The determination of nitrogen-15 by emission and mass spectrometry in biochemical analysis: a review

Methods for the establishment of 15N 14N ratios are reviewed. Mass spectrometric and emission spectrometric procedures are discussed in detail with particular reference to the analysis of biological materials. Some aspects of 15N chemistry are also described.

Variation of 15N in Corn and Soil Following Application of Fertilizer Nitrogen

AbstractSamples of corn (Zea mays L.) plant tissue (grain and leaf) were obtained from the University of Illinois South Farm plots and from the University of Illinois Morrow plots. Soil samples were also obtained from each of the South Farm plots. The 15N:14N ratio of the nitrogen in these corn plants is shown to be sensitive to the source of nitrogen taken up by the plants. A systematic decline in δ15N (per mill excess 15N) was observed within a narrow range of variation (about 4 δ15N units) with increasing applications of commercial nitrogen fertilizer. This result is consistent with increasing contributions of fertilizer nitrogen to the plant, as the fertilizer had a δ15N value close to zero. Similarly, the influence of a small amount of manure nitrogen (40 kg nitrogen/ha) applied over many years was detected in samples grown on the University of Illinois' historic Morrow plots. The results indicate that the nitrogen of grain and the leaf tissue of young plants is derived at least in part from different pools reflecting different proportions of fertilizer and soil nitrogen or fertilizer nitrogen of different δ15N, due to the time course of its fractionation. Unexplained differences between the influence of soybeans and clover on δ15N were also observed. Neither the quantity nor the δ15N of the total nitrogen was influenced by short term (3 years) addition of large quantities of nitrogen (448 kg/ha) to the soil on the South Farm plots.A relatively high value of δ15N was obtained for NO3‐ in the soil profile in the fall following spring application of 448 kg nitrogen/ha. This suggests that the observed nitrate nitrogen was not derived from nitrogen molecules applied as fertilizer in the spring, but originated rather from the exchange of fertilizer nitrogen for soil nitrogen and/or the “priming effect” (release of extra soil nitrogen as a result of the application of fertilizer nitrogen).

Nucleoside Antibiotics: EPIMERIZATION OF CARBON 2′ OF ADENOSINE DURING THE BIOSYNTHESIS OF 9-β-d-ARABINOFURANOSYLADENINE BY STREPTOMYCES ANTIBIOTICUS

Abstract By a combination of 15N-, 3H-, and 14C-labeled experiments with adenosine and d-ribose, it has been possible to demonstrate that adenosine is directly involved in the biosynthesis of 9-β-d-arabinofuranosyladenine by Streptomyces antibioticus. This conclusion is based on the following observations. (a) The ratio of 15N:14C in [6-amino-15N, U-14C]arabinofuranosyladenine and in the adenosine isolated from the RNA are the same as the ratio of 15N:14C in [6-amino-15N, U-14C]adenosine added to cultures of S. antibioticus. (b) All of the tritium on C-2' of [2'-3H]adenosine is lost in the epimerization. (c) Arabinofuranosyladenine derived from d-[3-3H]ribose contained only one-third of the tritium label on C-3'. (d) The conversion of adenosine to 9-β-d-arabinofuranosyladenine does not proceed via 2'-deoxyadenosine. (e) [U-14C]Arabinofuranosyladenine is rapidly taken up by S. antibioticus, but it is hydrolyzed to adenine. This latter observation was determined by the isolation of 14C-labeled AMP from the cellular RNA. These data suggest that adenosine is converted to a 2'-ketonucleoside or 2'-ketonucleotide. Enolization of this 2'-keto compound might explain the loss of tritium on the 3' carbon of arabinofuranosyladenine. The metabolism of 9-β-d-arabinofuranosyladenine and 2'-deoxyadenosine differs markedly from that of adenosine in that the first two nucleosides are hydrolyzed to adenine while adenosine is not hydrolyzed. If the conversion of adenosine to arabinofuranosyladenine proceeds via an epimerase, this reaction would differ from the UDP-d-glucose 4'-epimerase and l-ribulose-phosphate 4'-epimerase in which the hydrogen on C-4' of UDP-glucose or d-xylulose 5-phosphate is found intact in the same position following epimerization. The biosynthesis of arabinofuranosyladenine could be similar to the UDP-acetyl-d-glucosamine 2'-epimerase, cellobiose-2'-epimerase, d-ribulose-phosphate 3'-epimerase, or dTDP-l-rhamnose synthetase in which hydrogen from water is incorporated into the product.

Studies on Nitrogen Metabolism of Soybean Plants : II. Proportions of labelled and nonlabelled nitrogen transported to various parts of the plants

Transport of newly absorbed 15N and previously accumulated 14N in young soybean plants was examined following the application of nitrate- 15N to the root medium. During a 12-hour absorption period the roots exported both 15N and 14N to the shoots in the ratio of 51 : 49 for Experiment 1 and 82 : 18 for Experiment 2. A higher 15N ratio found in the Experiment 2 may be attributed to the aerated condition of the root medium, while in the Experiment 1 this condition was omitted. The proportion of 15N and 14N in the nitrogen transported to apical growing region of the stem showed a marked difference from that of the nitrogen exported from the roots with a considerably lesser 15N ratio to 14N such as 16 : 84 for Experiment 1 and 30 : 70 for Experiment 2. The total amount of nitrogen entering leaves during the experiment was estimated from the leaves with the "ringing" treatment to their petioles. It was found that the nitrogen transported into leaves also consisted of 15N and 14N components, and their ratios were intermediate between those of the roots and those of the apical growing regions. When comparison was made among the leaves of different ages, the percentage of 14N increased as the age decreased. Thus, it may be said that the relative content of 14N in the translocating nitrogen increased as the nitrogen moved through the stem toward the apex.

Biosynthesis of chloramphenicol. 3. Phenylpropanoid intermediates.

L-p-Aminophenylalanine-α-14C:α-15N was shown to be a specific precursor of the p-nitrophenylserinol moiety of chloramphenicol produced by cultures of Streptomyces species. It was incorporated with only a small change in the 14C:15N ratio. Evidence was obtained for the presence of a small intracellular pool of p-aminophenylalanine. DL-threo-p-Aminophenylserine-carboxyl-14C and α-N-dichloroacetyl-Ls-threo-p-aminophenylserinol-hydroxymethyl-14C were also incorporated into the antibiotic without randomization of the label.