Sesbania spp., Aeschynomene indica and Crotalaria spp. are amide-exporters

Abstract

Extract

Leguminous plants consist of two groups, amide-exporting and ureide-exporting plants. The former legumes export a large fraction of fixed-N in the form of amides (asparagine and glutamine), and the latter legumes in the form of ureides (allantoic acid and allantoin). Another characteristic of the nodules is the enrichment in ¹⁵N. There are two types of legumes: one characterized by the enrichment with ¹⁵N in N₂-fixing nodules, in contrast to the other where the enrichment does not occur. The first investigation by Shearer et al. (1982) suggested that the nodules exporting fixed-N in the form of ureides were enriched in ¹⁵N unlike those exporting it in the form of amides. Soybeans, mungbean, and cowpea belong to the former group and groundnut, alfalfa, white clover to the latter. Although pea and faba bean were first classified into the latter group (Shearer et al. 1982), a recent investigation (Yoneyama 1988) showed that these nodules were also enriched in ¹⁵N.     

Root Carbon Enrichment Alleviates Ammonium Toxicity in Cucumber Plants

ABSTRACT

The addition of carbonates to a nutrient solution to alleviate ammonium (NH₄ ⁺) toxicity in hydroponically-grown cucumber (Cucumis sativus L.) plants was investigated. Stable isotopes [nitrogen (¹⁵N) and carbon (¹³C)] were used to assess the uptake of nitrogen [NH₄ ⁺ or nitrate (NO₃ ^?)] as well as carbon [bicarbonate (HCO₃ ^?)/carbonate (CO₃ ^2?)] by the roots. Ammonium as the sole N source at 5 mM decreased plant fresh weights compared to NO₃ ^?. However, at lower concentrations of NH₄ ⁺ (25% of 5 mM total N), growth was increased compared to NO₃ ^? alone. Inorganic C enrichment [calcium carbonate (CaCO₃)] of the nutrient solution increased the fresh weight of NH₄ ⁺ grown plants with up to 150% relative to control plants receiving calcium hydroxide [Ca(OH)₂] for pH regulation. Root ¹⁵N enrichment was lower in ¹⁵NH₄ ⁺ supplied plants compared to ¹⁵NO₃ ^?, while the ¹³C enrichment in leaves was increased by NH₄ ⁺ nutrition compared to NO₃ ^? or NH₄NO_3. The enhanced C capture was associated with high PEPCase activity in the roots. It is concluded that inorganic carbon enrichment of the root medium may alleviate NH₄ ⁺ toxicity via increased synthesis of C skeletons and, accordingly, increased capacity for NH₄ ⁺ assimilation and N export to the shoots.     

Isotope 15N enrichment of soil-ammonium N as a reference to estimate N2 fixation by stem and root-inoculated S. rostrata

Summary A pot experiment in the greenhouse was conducted to compare the contribution of N derived from the atmosphere or from biological N₂ fixation by Sesbania rostrata inoculated with Azorhizobium caulinodans, applied either to roots or to roots and stems (single or multiple stem inoculation). Two subsequent crops were grown for 50 days under flooded conditions. N derived from air was estimated by ¹⁵N dilution using ¹⁵N enrichment of soil NH _inf4 ^sup+ -N and of Echinochloa crusgalli as the non-N₂-fixing reference datum and compared with estimates obtained by the N-difference method. The first crop was grown to stabilize the ¹⁵N into the soil organic N fraction. The ¹⁵N enrichment of soil NH _inf4 ^sup+ -N in the second crop declined slowly. The extractability ratio (¹⁵N enrichment of extractable soil N to ¹⁵N enrichment of total soil N) decreased from 4.8 to 4.1 50 days after planting. The enrichment of soil NH _inf4 ^sup+ -N was comparable to that of E. crus-galli, resulting in similar estimates of N derived from air when either soil NH _inf4 ^sup+ -N or enrichment of E. crus-galli was used as a non-fixing reference. The N-difference method did not always provide reliable estimates of N derived from air; percentages ranged from 75 to more than 80 by 50 days after planting in both crops and did not differ among treatments. The study demonstrates the potential of using ¹⁵N enrichment of soil NH _inf4 ^sup+ -N as a non-N₂-fixing reference for reliable BNF estimates of crops in lowland puddled soil.     

Estimating N2 fixation by field-grown lupins (Lupinus angustifolius L.) using soil and plant 15N enrichment

Summary Biological N₂ fixation was estimated in a field experiment following the addition of NH₄Cl or KNO₃ to unconfined microplots (1.5 m²) at 2.5 g N m^-2 (10 atom% ¹⁵N). A model of total N and ¹⁵N accumulation in lupins and decreasing ¹⁵N enrichment in the KCl-extractable soil-N pool (0–0.15 m depth) was used to estimate the proportion of N in lupins derived from biological N₂ fixation. Estimates of N₂ fixation derived from the model were compared with ¹⁵N isotope-dilution estimates obtained using canola, annual ryegrass, and wheat as nonfixing reference plants. Biomass, total N accumulation, or ¹⁵N enrichment in the lupin and reference crops did not differ whether NH _inf4 ^sup+ or NO _inf3 ^sup- was added as the labelled inorganic-N source. The decrease in soil ¹⁵N enrichment was described by first-order kinetics, whereas total N and ¹⁵N accumulation in the lupins were described by logistical equations. Using these equations, the uptake of soil N by lupins was estimated and was then used to calculate fixed N₂. Estimates of N₂ fixation derived from the model increased from 0 at 50 days after sowing to a maximum of 0.79 at 190 days after sowing. Those based on the ¹⁵N enrichment of the NO _inf3 ^sup- pool were 10% higher than those based on the mineral-N pool. ¹⁵N isotope-dilution estimates of N₂ fixation ranged from 0.37 to 0.55 at 68 days after sowing and from 0.71 to 0.77 at 190 days after sowing. Reference plant-derived values of N₂ fixation were all higher than modelled estimates during the early states of growth, but were similar to modelled estimates at physiological maturity. The use of the model to estimate N₂ derived from the atmosphere has the intrinsic advantage that the need for a non-fixing reference plant is avoided.     

Effects of Multiple Reference Plants,Season, and Irrigation on Biological Nitrogen Fixation by Pasture Legumes using the Isotope Dilution Method

Abstract

The popular and widely used ¹⁵nitrogen (N)–isotope dilution method for estimating biological N fixation (BNF) of pasture and tree legumes relies largely on the ability to overcome the principal source of error due to the problem of selecting appropriate reference plants. A field experiment was conducted to evaluate the suitability of 12 non‐N₂‐fixing plants (i.e., nonlegumes) as reference plants for estimating the BNF of three pasture legumes (white clover, Trifolium repens L.; lucerne, Medicago sativa; and red clover, Trifolium pratense L.) in standard ryegrass–white clover (RWC) and multispecies pastures (MSP) under dry‐land and irrigation systems, over four seasons in Canterbury, New Zealand. The ¹⁵N‐isotope dilution method involving field ¹⁵N‐microplots was used to estimate BNF. Non‐N₂‐fixing plants were used either singly or in combination as reference plants to estimate the BNF of the three legumes. Results obtained showed that, on the whole, ¹⁵N‐enrichment values of legumes and nonlegumes varied significantly according to plant species, season, and irrigation. Grasses and herb species showed higher ¹⁵N‐enrichment than those of legumes. Highest ¹⁵N‐enrichment values of all plants occurred during late summer under dry‐land and irrigation conditions. Based on single or combined non‐N₂‐fixing plants as reference plants, the proportion of N derived from the atmosphere (% Ndfa) values were high (50 to 90%) and differed between most reference plants in the MSP pastures, especially chicory (Cichorium intybus), probably because it is different in phenology, rooting depth, and N‐uptake patterns compared to those of legumes. The percent Ndfa values of all plants studied also varied according to plant species, season, and irrigation in the MSP pastures. Estimated daily amounts of BNF varied according to pasture type, time of plant harvest, and irrigation, similar to those shown by percent Ndfa results as expected. Irrigation increased daily BNF more than 10‐fold, probably due to increased dry‐matter yield of pasture under irrigation compared to dry‐land conditions. Seasonal and irrigation effects were more important in affecting estimates of legume BNF than those due to the appropriate matching of N₂‐fixing and non‐N₂‐fixing reference plants.     

Estimating N2 fixation by Sesbania rostrata and S. cannabina (syn. S. aculeata) in lowland rice soil by the 15N dilution method

Summary A field experiment in concrete-based plots was conducted to estimate the contribution of N derived from air (Ndfa) or biological N₂ fixation in Sesbania rostrata and S. cannabina (syn. S. aculeata), using various references, by the ¹⁵N dilution method. The two Sesbania species as N₂-fixing reference plants and four aquatic weed species as non-N₂-fixing references were grown for 65 days after sowing in two consecutive crops, in the dry and the wet seasons, under flooded conditions. Soil previously labeled with ¹⁵N at 0.26 atom % ¹⁵N excess in mineralizable N was further labeled by ammonium sulfate with 3 and 6 atom % ¹⁵N excess. The results showed that ¹⁵N enrichment of soil NH ₄ ⁺ -N dropped exponentially in the first crop to half the original level in 50 days while in the second crop, it declined gradually to half the level in 130 days. The decline in ¹⁵N enrichment, in both N₂-fixing and non-fixing species, was also steeper in the first crop than in the second crop. Variations in ¹⁵N enrichment among non-fixing species were smaller in the second crop. The ratio of the uptake of soil N to that of fertilizer N in N₂-fixing and non-fixing species was estimated by the technique of varying the ¹⁵N level. In the second crop, this ratio in non-fixing species was higher than that in N₂-fixing species. Comparable estimates of % Ndfa were obtained by using ¹⁵N enrichment of various non-fixing species. There was also good agreement between the estimates obtained by using ¹⁵N enrichment of non-fixing species and those by using soil NH ₄ ⁺ -N, particularly in the second crop. By 25 days after sowing, the first crop of both Sesbania spp. had obtained 50% of total N from the atmosphere and the second crop had obtained 75%. The contribution from air increased with the age of the plant and ranged from 70% to 95% in 45–55 days. S. rostrata fixed substantially higher amounts of N₂ due to its higher biomass production compared with S. cannabina. Mathematical considerations in applying the ¹⁵N dilution method are discussed with reference to these results.     

Ammonium immobilisation during chloroform fumigation

The assumption in using the chloroform fumigation technique for microbial biomass determination is that microbes are killed or at least inactivated by the treatment. Problems associated with transformations of the N released on or during fumigation have so far only been associated with the fumigation-incubation method. A laboratory and a field study were carried out to investigate the possible N transformations during biomass determination by the fumigation-extraction method. Labelled NH₄NO₃ (either the NO₃⁻, NH₄⁺ or both pools were ¹⁵N enriched) was applied to the soil and biomass determinations made at intervals subsequently. The size and enrichment of the ammonium (NH₄⁺), and nitrate (NO₃⁻) pools were determined before and after chloroform fumigation. The ¹⁵N enrichment of the NH₄⁺ pool after fumigation could only be explained if immobilisation of ammonium occurred at some time during the 24 h fumigation period. The extent of this immobilisation was calculated. In addition, there was evidence that nitrification occurred during the fumigation procedure at the start of the laboratory study and throughout the field study. The laboratory and field study differed mainly in the dynamics related to NO₃⁻ uptake and release. There was evidence for uptake of NO₃⁻ by the microbial biomass with and without utilization. We conclude that the ¹⁵N enrichment in the microbial biomass cannot be accurately determined when N transformations and release of non-utilized N occurs during fumigation. The possible immobilisation of mineral N during fumigation will affect the magnitude of the factor used to convert measured microbial biomass N to actual microbial biomass N in soil.     

Elevated CO2 concentrations increase leaf nitrate reduction by strengthening sink activity in soybean plants

An experiment was conducted to examine the effect of CO₂ enrichment on the nitrate uptake, nitrate reduction activity, and translocation of assimilated-N from leaves at varying levels of nitrogen nutrition in soybean using ¹⁵N tracer technique. CO₂ enrichment significantly increased the plant biomass, apparent leaf photosynthesis, sugar and starch contents of leaves, and reduced-N contents of the plant organs only when the plants were grown at high levels of nitrogen. A high supply of nitrogen enhanced plant growth and increased the reduced-N content of the plant organs, but its effect on the carbohydrate contents and photosynthetic rate were not significant. However, the combination of high CO₂ and high nitrogen levels led to an additive effect on all these parameters. The nitrate reductase activity increased temporarily for a short period of time by CO₂ enrichment and high nitrogen levels. ¹⁵N tracer studies indicated that the increase in the amount of reduced-N by CO₂ enrichment was derived from nitrate-N and not from fixed-N of the plant. To examine the translocation of reduced-N from the leaf in more detail, another experiment was conducted by feeding the plants with ¹⁵NO₃-N through a terminal leaflet of an upper trifoliated leaf under depodding and/or CO₂ enrichment conditions. The export rate of ¹⁵N from the terminal leaflet to other plant parts decreased by depodding, but it increased by CO₂ enrichment. CO₂ enrichment increased the percentage of plant ¹⁵N in the stem and / or pods. Depodding increased the percentage of plant ¹⁵N in the leaf and stem. The results suggested that the increase in the leaf nitrate reduction activity by CO₂ enrichment was due to the increase of the translocation of reduced-N from leaves through the strengthening of the sink activity of pods and / or stem for reduced-N.     

Incomplete Acetylene Inhibition of Nitrous Oxide Reduction in Potential Denitrification Assay as Revealed by using 15N-Nitrate Tracer

One lake sediment and three soils for rice production were used to test the effectiveness of inhibiting of nitrous oxide (N₂O) reduction to dinitrogen gas (N₂) by acetylene (C₂H₂) using ¹⁵N tracer. Regardless of the sources of the samples, results show that in presence of C₂H₂, significant isotopic enrichment of ¹⁵N of N₂ was found at end of a typical denitrification assay. The δ¹⁵N of N₂ value increased from 0‰ to 7.8–19.3‰ and 7.5–10.6‰ for the treatment with addition of 0.05 and 0.2 mg ¹⁵N nitrate, respectively. Such ¹⁵N enrichment can be interpreted as N₂ formation accounting for 15.3% and 2.5% of the total added N in these two treatments, respectively. Nitrous oxide accumulation in presence of C₂H₂ could not account for the total added N. The result indicates incomplete inhibition of N₂O reduction to N₂ by C₂H₂ in denitrification when N₂O reduction enzyme is developed.     

Technique of soil sample pretreatment for analysis of 15N:14N isotope ratio

Abstract

The technique of simultaneous quantitative determination of mineral N soil forms (nitrates, exchangeable and non‐exchangeable ammonium, and total amount of these compounds) and sample pretreatment for the analysis of ¹⁵N:¹⁴N ratio is suggested. The technique is based on the selective association of NH₄ ⁺‐ions into indophenol complex and subsequent ethyl‐acetate extraction of this complex from solution. The mineralization of indophenol is carried out in alkaline medium with simultaneous NH₃ distillation into H₂SO₄ titrant. The application of given technique allows us to shorten significantly the time required for analysis and to increase the accuracy of analytical determination.     

Temporal responses of soil microorganisms to substrate addition as indicated by amino sugar differentiation

Amino sugars are one of the important microbial residue biomarkers which are associated with soil organic matter cycling. However, little is known about their transformation kinetics in response to available substrates because living biomass only contributes a negligible portion to the total mass of amino sugars. By using ¹⁵N tracing technique, the newly synthesized (labeled) amino sugars can be differentiated from the native portions in soil matrix, making it possible to evaluate, in quantitative manner, the transformation pattern of amino sugars and to interpret the past and ongoing changes of microbial communities during the assimilation of extraneous ¹⁵N. In this study, laboratory incubations of soil samples were conducted by using ¹⁵NH₄⁺ as nitrogen source with or without glucose addition. Both the ¹⁵N enrichment (expressed as atom percentage excess, APE) and the contents of amino sugars were determined by an isotope-based gas chromatography-mass spectrometry. The significant ¹⁵N incorporation into amino sugars was only observed in glucose plus ¹⁵NH₄⁺ amendment with the APE arranged as: muramic acid (MurN) > glucosamine (GlcN) > galactosamine (GalN). The dynamics of ¹⁵N enrichment in bacterial-derived MurN and fungal-derived GlcN were fitted to the hyperbolic equations and indicative for the temporal responses of different soil microorganisms. The APE plateau of MurN and fungal-derived GlcN represented the maximal extent of bacterial and fungal populations, respectively, becoming active in response to the available substrates. The different dynamics of the ¹⁵N enrichment between MurN and GlcN indicated that bacteria reacted faster than fungi to assimilate the labile substrates initially, but fungus growth was dominant afterward, leading to integrated microbial community structure over time. Furthermore, the dynamics of labeled and unlabeled portions of amino sugars were compound-specific and substrate-dependent, suggesting their different stability in soil. GlcN tended to accumulate in soil while MurN was more likely degraded as a carbon source when nitrogen supply was excessive.     

Arbuscular mycorrhizal fungi contribute to C and N enrichment of soil organic matter in forest soils

Increasing evidence suggests that accretion of microbial turnover products is an important driver for isotopic carbon (C) and nitrogen (N) enrichment of soil organic matter (SOM). However, the exact contribution of arbuscular mycorrhizal fungi (AMF) to soil isotopic patterns remains unknown. In this study, we compared ¹³C and ¹⁵N patterns of glomalin-related soil protein (GRSP), which includes a main fraction derived from AMF, litter, and bulk soil in four temperate rainforests. GRSP was an abundant C and N pool in these forest soils, showing significant ¹³C and ¹⁵N enrichment relative to litter and bulk soil. Hence, cumulative accumulation of recalcitrant AMF turnover products in the soil profile likely contributes to ¹³C and ¹⁵N enrichment in forest soils. Further research on the relationship between GRSP and AMF should clarify the exact extent of this process.     

Effect of successive cuttings on uptake and partitioning of 15N among plant parts of Leucaena leucocephala

Summary We studied the effect of three successive cuttings on N uptake and fixation and N distribution in Leucaena leucocephala. Two isolines, uninoculated or inoculated with three different Rhizobium strains, were grown for 36 weeks and cut every 12 weeks. The soil was labelled with 50 ppm KNO₃ enriched with 10 atom % ¹⁵N excess soon after the first cutting. Except for the atom % ¹⁵N excess in branches of K28 at the second cutting, both the L. leucocephala isolines showed similar patterns of total N, fixed N₂, and N from fertilizer distribution in different parts of the plant at each cutting. The Rhizobium strain did not influence the partitioning of ¹⁵N among the different plant parts. Significant differences in ¹⁵N enrichment occurred in different parts. Live nodules of both isolines showed the lowest atom % ¹⁵N excess values (0.087), followed by leaves (0.492), branches (0.552), stems (0.591), and roots (0.857). The roots contained about 60% of the total plant N and about 70% of the total N derived from fertilizer over the successive cuttings. The total N₂ fixed in the roots was about 60% of that fixed in the whole plant, while the shoots contained only 20% of the fixed N₂. We conclude that N reserves in roots and nodules constitute another N source that must be taken into account when estimating fixed N₂ or the N balance after pruning or cutting plants. ¹⁵N enrichment declined up to about fivefold in the reference and the N₂-fixing plants over 24 weeks following the ¹⁵N application. The proportion and the amounts of N derived from fertilizer decreased, while the amount derived from N₂ fixation increased with time although its proportion remained constant.     

应用15N稀释法测定土壤氮素初级转化速率的一些关键技术

本文综合评述了应用~(15)N库稀释法测定土壤氮素初级转化速率的一些关键技术,即~(15)N标记土壤氮库的方法、~(15)N的加入量、丰度和标记物种类的选择,以及初始取样时间的确定。只有合理地运用这些关键技术,才能更准确地测定土壤氮素初级转化速率,进而更真实地表征土壤氮素的实际周转状况。     

The difference in N metabolism between NAD(P)H-specific NR-deficient mutant and wild-type barley (Hordeum vulgare L.)

Abstract

Barley (Hordeum vulgare L.) is an important crop for cereal research. In this study, two barley genotypes the wild-type (Steptoe) and the mutant (Az12) were used. An experiment was conducted using ¹⁵N-tracing method to NADH-specific nitrate reductase (NR)-deficient mutant seedling of barley. The N-depleted seedlings were exposed to a nutrient solution containing nitrate and nitrite, and were labeled with ¹⁵N for 38?h under (14?L/10D) cycles. The two genotypes utilized ¹⁵NO₃^? and accumulated it as reduced ¹⁵N, predominately in the shoots. However, nitrate reduction in the Az12 shoots was 9% lower than that in the Steptoe shoots at 38?h. As a result, in the Az12, nitrate accumulation in shoots was 78% higher than that in the Steptoe. Accumulation of reduced ¹⁵N in the Az12 roots was nearly similar to that of the Steptoe roots, but 8% lower in the Az12 shoots than in the Steptoe shoots at the end of the experiment. Also for both genotypes, root contribution increased during L/D cycles and decreased during the subsequent light cycle. Upward transport of reduced ¹⁵N via the xylem in the Az12 was nearly two times higher than that in Steptoe during the second light period (24–38?h). In both genotypes, xylem transport of reduced ¹⁵N was far exceeded the downward phloem transport. Abbreviations Anl accumulation of reduced ¹⁵N from ¹⁵NO₃^? in non-labeled roots of split roots

Ar accumulation in roots of reduced ¹⁵N from ¹⁵NO₃^?

As accumulation in shoots of reduced ¹⁵N from ¹⁵NO₃^?

Rr ¹⁵NO₃^? reduction in roots

Rs ¹⁵NO₃^? reduction in shoots

Tp translocation to root of shoot reduced ¹⁵N from ¹⁵NO₃^? in phloem

Tx translocation to shoot of root-reduced ¹⁵N from ¹⁵NO₃^? in xylem

FW fresh weight

     

Evidence of carbon stimulated N transformations in grassland soil after slurry application

High nitrification rates which convert ammonium (NH₄⁺) to the mobile ions NO₂⁻ and NO₃⁻ are of high ecological significance because they increase the potential for N losses via leaching and denitrification. Nitrification can be performed by chemoautotrophic or heterotrophic organisms and heterotrophic nitrifiers can oxidise either mineral (NH₄⁺) or organic N. Selective nitrification inhibitors and ¹⁵N tracer studies have been used in an attempt to separate heterotrophic and autotrophic nitrification. In a laboratory study we determined the effect of cattle slurry on the oxidation of mineral NH₄⁺-N and organic-N by labelling the NH₄⁺ or NO₃⁻ pools separately or both together with ¹⁵N. The size and enrichment of the mineral N pools were determined at intervals. To calculate gross N transformation rates a ¹⁵N tracing model was developed. This model consists of the three N-pools NH₄⁺, NO₃⁻ and organic N. Sub-models for decomposition of degradable carbon in the soil and the slurry were added to the model and linked to the N transformation rates. The model was set up in the software ModelMaker which contains non-linear optimization routines to determine model parameters. The application of cattle slurry increased the rate of nitrifcation by a factor of 20 compared with the control. The size and enrichment of the mineral N pools provided evidence that nitrification was due to the conversion of NH₄⁺ to NO₃⁻ and not the conversion of organic N to NO₃⁻. There was evidence that slurry-enhanced oxidation of NH₄⁺ to NO₃⁻ was due to a combination of autotrophic and heterotrophic transformations. Slurry application increased the mineralisation rate by approximately a factor of two compared with the control and the rate of immobilisation of NH₄⁺ by approximately a factor of three.     

Fate of Fertilizer-15N to a Maize Crop Grown in Northern Zambia

Abstract

A field study with maize (Zea mays L.) was conducted in the 1988/89 cropping season to investigate the fate of ¹⁵NO₃-N-labelled NH₄ ¹⁵NO₃ applied at 40, 80 and 120 kg N ha^?1 (unlabelled N applied at 0, 80, 160 and 240 N ha^?1) with and without lime. The investigations were conducted in northern Zambia at Misamfu Regional Research Centre, Kasama on a Misamfu red sandy loam soil. The experimental design was a split plot arrangement with four replications with main plots receiving 0 and 2 Mg ha^?1 dolomitic limestone, while subplots received fertilizer N at various rates. Significant (p < 0.001) grain and DM yield responses to applied N up to 160 kg ha^?1 were observed. At higher rates little or no crop responses were observed and fertilizer use efficiency declined. Partitioning of amounts of total N and ¹⁵N in plants was in the order of seed = tassel > leaf> cob = earleaf> stem. Fertilizer N rates showed a highly significant (p < 0.001) effect on plant uptake of labelled N. Lime and its interaction with N rates had no effect on all measured parameters. Leaching of NO₃-N fertilizer to lower soil depths was in proportion to the rate of N applied, with highly significant (p < 0.001) differences among soil depths. Although higher concentrations of fertilizer-¹⁵N were recovered in the 0–20 cm depth the recovered portion at lower soil depths was still significant. Total recovery of labelled N by plant and by soil after crop harvest averaged 75, 55 and 54% of originally applied fertilizer-¹⁵N at 40, 80 and 120 kg N ha^?1, respectively. Corresponding unaccounted for ¹⁵N was 25, 45 and 46%. The most probable loss mechanism could have been by leaching to depths greater than 60 cm, gaseous losses to the atmosphere and root assimilation.     

A GC/MS method for the assessment of N and C incorporation into soil amino acid enantiomers

Identifying the transformation process of amino acid enantiomers was essential to probe into the fate, turnover and aging of soil nitrogen due to their important roles in the biogeochemical cycling. If this can be achieved by differentiating between the newly biosynthesized and the inherent compounds in soil, then the isotope tracer method can be considered most valid. We thereby developed a gas chromatography/mass spectrometry (GC/MS) method to trace the ¹⁵N or ¹³C isotope incorporation into soil amino acid enantiomers after being incubated with ¹⁵NH₄⁺ or U-¹³C-glucose substrates. The most significant fragments (F) as well as the related minor ions were monitored by the full scan mode and the isotope enrichment in amino acids was estimated by calculating the atom percentage excess (APE). ¹⁵NH₄⁺ incorporation was evaluated according to the relative abundance increase of m/z F+1 to F for neutral and acidic amino acids and F+2 to F (mass 439) for lysine. The assessment of ¹³C enrichment in soil amino acids was more complicated than that of ¹⁵N due to multi-carbon atoms in amino acid molecules. The abundance ratio increment of m/z F+n to F (n is the original skeleton carbon number in each fragment) indicated the direct conversion from the added glucose to amino acids, but the total isotope incorporation from the added ¹³C can only be calculated according to all target isotope fragments, i.e. the abundance ratio increment summation from m/z (Fa+1) through m/z (Fa+T) represented the total incorporation of the added ¹³C (Fa is the fragment containing all original skeleton carbons and T is the carbon number in the amino acid molecule). This method has a great advantage especially for the evaluation of high-abundance isotope enrichment in organic compounds compared with GC/C/IRMS. And in principle, this technique is also valid for amino acids besides enantiomers if stereoisomers are not concerned. Our assessment approach could shine a light on investigating the biochemical mechanism of microbial transformation of N and C in soils of terrestrial ecosystem.     

Short‐Term Recovery of Ammonium‐15Nitrogen Applied to a Temperate Forest Inceptisol and Ultisol in East Tennessee,USA

Abstract

The short‐term fate and retention of ammonium (NH₄)‐¹⁵nitrogen (N) applied to two types of forest soils in east Tennessee was investigated. Four ridgetop forests, predominantly oak (Quercus spp.), were studied. Five applications of NH₄‐¹⁵N tracer were made to the forest floor at 2‐ to 4‐week intervals over a 14‐week period in 2004. Nitrogen‐15 recovery in the forest floor, fine roots (<2 mm), and the mineral soil (0–20 cm) was calculated at 6, 21, and 42 weeks after the last application. Most of the ¹⁵N was retained in the forest floor and the mineral soil, with only small amounts (≤2%) found in roots from both soil layers. Recovery of NH₄‐¹⁵N was greater in Inceptisols, which had a wider carbon (C)‐to‐N ratio than Ultisols. For both soil types, higher NH₄‐¹⁵N recoveries and long retention times (half‐lives>100 weeks) indicated the forest floor is an effective filter for atmospheric N inputs.     

A method to assess whether ‘preferential use’ occurs after N ammonium addition; implication for the N isotope dilution technique

The robustness of the assumption of equilibrium between native and added N during ¹⁵N isotope dilution has recently been questioned by Watson et al. (Soil Biol Biochem 32 (2000) 2019-2030). We re-analyzed their raw data using equations that consider the added and native NH₄⁺ and NO₃⁻ pools as separate state variables. Gross mineralization rates and first-order rate constants for NH₄⁺ and NO₃⁻ consumption were obtained by combining analytical integration of the differential equations with a non-linear fitting procedure. The first-order rate constants for NH₄⁺ consumption and NO₃⁻ immobilization for the added NH₄⁺ and NO₃⁻ pool were used to estimate gross mineralization rates and first-order rate constants for nitrification of native NH₄⁺. The latter were 2-4 times lower than the first-order rate constants derived from the added N pool. This discrepancy between first-order rate constants for nitrification implies that one or more process rates estimated for the added N pools cannot be applied to the native N pools. Preferential use of the added N resulted in an overestimation of the gross mineralization by 1.5-2.5-fold, emphasizing the need for critical evaluation of the assumption of equilibrium before gross mineralization rates are calculated.