Nitrogen isotope analysis of free glycine in soil using isotope dilution mass spectrometry

To enable the estimation of production and consumption rates of free glycine in soils through ¹⁵N isotope dilution experiments, an isotope dilution mass spectrometric method was developed. The method, which enabled high precision N isotope ratio determination of glycine in soil extracts at δ¹⁵N levels up to 4000‰ and concentrations from approximately 2 μM, is based on the following steps: (i) addition of glycine spike to the soil extract, (ii) removal of humic substances and pre-concentration of glycine using solid phase extraction, (iii) derivatization of amino acids, (iv) separation of the derivatives using gas chromatography (GC), (v) their combustion to yield sample N₂ gas, and (vi) finally the use of N isotope ratio mass spectrometry (IRMS). Judging by uncertainty budget calculations, the precision obtained (SD=0.01-0.06 at% ¹⁵N) is sufficient for detecting differences in N isotopic ratios obtained in ¹⁵N isotope dilution experiments.     

铬污染归宿的同位素原位监测法

铬(Cr)污染的迁移和环境毒性主要取决于其化学形态。在环境中,如果移动性及毒性较大的Cr(Ⅵ)能被还原为移动性及毒性较小的Cr(Ⅲ),并且还原速率大于污染速率,那么对铬污染的自然生态修复治理是可行的。铬在氧化还原过程中,存在着同位素动力学分馏效应,致使同位素比率发生移动,移动的大小和反应进程有关,服从Reyleigh关系方程。因此,利用同位素测定可以确定Cr(Ⅵ)的归宿,用于铬(Cr)污染自然修复的监测。本文就环境铬污染自然生态修复中,应用同位素原位分析法进行监测的基本原理、分析过程及应用的可行性进行了阐述。     

Impacts of anthropogenic N additions on nitrogen mineralization from plant litter in exotic annual grasslands

Urban regions of southern California receive up to 45 kg N ha^-1 y^-1 from nitrogen (N) deposition. A field decomposition study was done using ¹⁵N-labelled litter of the widespread exotic annual grass Bromus diandrus to determine whether elevated soil N is strictly from N deposition or whether N mineralization rates from litter are also increased under N deposition. Tissue N and lignin concentrations, which are inversely related in field sites with high and low N deposition, determine the rate at which N moves from plant litter to soil and becomes available to plants. The effect of soil N on N movement from litter to soil was tested by placing litter on high and low N soil in a factorial experiment with two levels of litter N and two levels of soil N. The litter quality changes associated with N deposition resulted in faster rates of N cycling from litter to soil. Concentrations of litter-derived N in total N, NH₄⁺, NO₃⁻, microbial N and organic N were all higher from high N/low lignin litter than from low N/high lignin litter. Litter contributed more N to soil NH₄⁺ and microbial N in high N than low N soil. At the end of the study, N mineralized from high N litter on high N soil accounted for 46% of soil NH₄⁺ and 11% of soil NO₃⁻, compared to 35% of soil NH₄⁺ and 6% of soil NO₃⁻ from low N litter on low N soil. The study showed that in high N deposition areas, elevated inorganic soil N concentrations at the end of the summer N deposition season are a result of N mineralized from plant litter as well as from N deposition.     

A survey of symbiotic nitrogen fixation by white clover grown on metal contaminated soils

There is conflicting evidence about toxic effects of heavy metals in soil on symbiotic nitrogen fixation. This study was set-up to assess the general occurrence of such effects. Soils with metal concentration gradients were sampled from six established field trials, where sewage sludge or metal salts have been applied, or from a transect in a sludge treated soil. Additional contaminated soils were sampled near metal smelters, in floodplains, in sludge amended arable land and in a metalliferous area. Symbiotic nitrogen fixation was measured with ¹⁵N isotope dilution in white clover (Trifolium repens L.) grown in potted soil that was not re-inoculated, and using ryegrass (Lolium perenne L.) as reference crop. The fraction nitrogen in clover derived from fixation (N_dff) varied from 0 to 88% depending on soil. Pronounced metal toxicity on N_dff was only confirmed in a sludge treated soil where nitrogen fixation was halved from the control value at soil total metal concentration of 737 mg Zn kg⁻¹, 428 mg Cu kg⁻¹ and 10 mg Cd kg⁻¹. The N_dff was significantly reduced by increasing metal concentration in soils from two other sites where N_dff was low throughout and where these effects might be attributed to confounding factors. No significant effects of metals on N_dff were identified in all other gradients even up to elevated total metal concentration (e.g. 55 mg Cd kg⁻¹). The variation of N_dff among all soils (n=48), is mainly explained by the number of rhizobia in the soil (log MPN, log (cells g⁻¹ soil)), whereas correlations with total or soil solution metal concentrations were weak (R²<0.25). The is significantly affected by the presence or absence of the host plant at the sampling site. No effects of metals were identified at even at total Zn concentrations of about 2000 mg Zn kg⁻¹, whereas metal toxicity could be identified at lower most probable number (MPN) values. This survey shows that the metal toxicity on symbiotic nitrogen fixation cannot be generalized and that survival of a healthy population of the microsymbiont is probably the critical factor.     

土壤容重对土壤供氮能力及棉花吸收利用氮素的影响

本文应用~(15)N示踪法研究不同土壤容重对土壤供氮能力及棉花吸氮的影响。阐明了丘陵黄棕壤地区棉花低产的原因是棉田土壤容重过大,使土壤的供氮能力——A_N值显著下降,棉株对氮素养分的吸收量减少,而且延缓了氮素养分由营养器官向生殖器官的转运。降低土壤容重是挖掘丘陵黄棕壤地区棉田增产潜力的有效施施。     

Nano-scale secondary ion mass spectrometry — A new analytical tool in biogeochemistry and soil ecology: A review article

Soils are structurally heterogeneous across a wide range of spatio-temporal scales. Consequently, external environmental conditions do not have a uniform effect throughout the soil, resulting in a large diversity of micro-habitats. It has been suggested that soil function can be studied without explicit consideration of such fine detail, but recent research has indicated that the micro-scale distribution of organisms may be of importance for a mechanistic understanding of many soil functions. Current techniques still lack the adequate sensitivity and resolution for data collection at the micro-scale, and the question ‘How important are various soil processes acting at different scales for ecological function?’ is therefore challenging to answer. The nano-scale secondary ion mass spectrometer (NanoSIMS) represents the latest generation of ion microprobes, which link high-resolution microscopy with isotopic analysis. The main advantage of NanoSIMS over other secondary ion mass spectrometers is its ability to operate at high mass resolution, whilst maintaining both excellent signal transmission and spatial resolution (down to 50 nm). NanoSIMS has been used previously in studies focussing on presolar materials from meteorites, in material science, biology, geology and mineralogy. Recently, the potential of NanoSIMS as a new tool in the study of biophysical interfaces in soils has been demonstrated. This paper describes the principles of NanoSIMS and discusses the potential of this tool to contribute to the field of biogeochemistry and soil ecology. Practical considerations (sample size and preparation, simultaneous collection of isotopes, mass resolution, isobaric interference and quantification of the isotopes of interest) are discussed. Adequate sample preparation, avoiding bias due to artefacts, and identification of regions-of-interest will be critical concerns if NanoSIMS is used as a new tool in biogeochemistry and soil ecology. Finally, we review the areas of research most likely to benefit from the high spatial and high mass resolution attainable with this new approach.     

Spatial variation of soil organic carbon concentrations and stable isotopic composition in 1-ha plots of forest and pasture in Costa Rica: implications for the natural abundance technique

The goal of this study was to examine spatial variation of soil organic C and its stable isotopic composition (δ¹³C) in 1-ha plots of mature rain forest and a cattle pasture dominated by C4 grasses in Costa Rica. Soil samples were taken from 80 mapped locations per plot and analyzed for organic C and δ¹³C. The range of values for soil C concentrations was similar between forest and pasture, although the mean values were higher in the forest. δ¹³C was narrowly constrained in the forest (from −27.96 to −26.09‰) but varied from −15.09 to −28.59‰ in the pasture. Variograms revealed spatial autocorrelation of soil C and δ¹³C in the pasture and organic C concentration in the forest soil. The large range and spatial variability of δ¹³C in the pasture site may be due to varying contributions of C3 and C4 vegetation to the soil C pool, which may limit the usefulness of the natural abundance technique as a precise tracer of soil C dynamics in this pasture.     

Effect of potassium phosphate fertilization on production and emission of methane and its C-stable isotope composition in rice microcosms

Rice fields are an important source for atmospheric CH₄, but the effects of fertilization are not well known. We studied the turnover of CH₄ in rice soil microcosms without and with addition of potassium phosphate. Height and tiller number of rice plants were higher in the fertilized than in the unfertilized microcosms. Emission rates of CH₄ were also higher, but porewater concentrations of CH₄ were lower. The δ¹³C values of the emitted CH₄ and of the CH₄ in the porewater were both 2-6% higher in the fertilized microcosms than in the control. Potassium phosphate did not affect rate and isotopic signature of CH₄ production in anoxic soil slurries. On the other hand, roots retrieved from fertilized microcosms at the end of incubation exhibited slightly higher CH₄ production rates and slightly higher CH₄-δ¹³C values compared to roots from unfertilized plants. Addition of potassium phosphate to excised rice roots generally inhibited CH₄ production and resulted in increasingly lower δ¹³C values of the produced CH₄. Fractionation of ¹³C during plant ventilation (i.e. δ¹³C in pore water CH₄ versus CH₄ emitted) was larger in the fertilized microcosms than in the control. Besides plant ventilation, this difference may also have been caused by CH₄ oxidation in the rhizosphere. However, calculation from the isotopic data showed that less than 27% of the produced CH₄ was oxidized. Collectively, our results indicate that potassium phosphate fertilization stimulated CH₄ emission by enhancing root methanogenesis, plant ventilation and/or CH₄ oxidation, resulting in residence times of CH₄ in the porewater in the order of hours.     

Spatial heterogeneity in the relocation of added C within the structure of an upland grassland soil

A pulse of ¹³CO₂ was added to the above ground vegetation in an upland grassland to determine the effects of faunal diversity on the flux of carbon to the surface horizons of the soil. Faunal diversity was manipulated by liming and biocide treatments for three years prior to the pulse addition. The relocation of ¹³C within roots and rhizosphere soil was determined by analysis of samples of bulk soil and of specific features identified on soil thin sections on four dates after the addition of the ¹³CO₂ pulse. Analysis of bulk soils showed only a small enrichment in ¹³C and no significant effects of the treatments. Analysis by isotope ratio mass spectrometry of the products of in situ laser combustion of root material and aggregates formed from faunal excrement showed that the distribution of the newly photosynthesised ¹³C is very localised, with large spatial variability in soil and root δ¹³C at scales of less than 1 mm. δ¹³C values ranged from the natural abundance level of around −28‰ to −4.9‰ in roots and to −8.4‰ in aggregates. The small pulse and large spatial variability masked any effects of the liming and biocide treatments in these soils. However, the variability in the relocation of newly photosynthesised carbon may help to explain the large spatial variability found in bacterial numbers at the sub-mm scale within soils and emphasises the importance of the accessibility of substrates to decomposers in undisturbed structured soils.     

Microbial immobilisation of C rhizodeposits in rhizosphere and root-free soil under continuous C labelling of oats

A greenhouse experiment was conducted by growing oats (Avenasativa L.) in a continuously ¹³CO₂ labeled atmosphere. The allocation of ¹³C-labeled photosynthates in plants, microbial biomass in rhizosphere and root-free soil, pools of soil organic C, and CO₂ emissions were examined over the plant's life cycle. To isolate rhizosphere from root-free soil, plant seedlings were placed into bags made of nylon monofilament screen tissue (16 μm mesh) filled with soil. Two peaks of ¹³C in rhizosphere pools of microbial biomass and dissolved organic carbon (DOC), as well as in CO₂ emissions at the earing and ripeness stages were revealed. These ¹³C maxima corresponded to: (i) the end of rapid root growth and (ii) beginning of root decomposition, respectively. The δ¹³C values of microbial biomass were higher than those of DOC and of soil organic matter (SOM). The microbial biomass C accounted for up to 56 and 39% of ¹³C recovered in the rhizosphere and root-free soil, respectively. Between 4 and 28% of ¹³C assimilated was recovered in the root-free soil. Depending on the phenological stage, the contribution of root-derived C to total CO₂ emission from soil varied from 61 to 92% of total CO₂ evolved, including 4-23% attributed to rhizomicrobial respiration. While 81-91% of C substrates used for microbial growth in the root-free soil and rhizosphere came from SOM, the remaining 9-19% of C substrates utilized by the microbial biomass was attributable to rhizodeposition. The use of continuous isotopic labelling and physical separation of root-free and rhizosphere soil, combined with natural ¹³C abundance were effective in gaining new insight on soil and rhizosphere C-cycling.