Impact of top-pruning time on the fertilizer N use efficiency of flue-cured tobacco as assessed by 15N tracing technique

To evaluate the impact of top-pruning time on fertilizer N use efficiency (NUE) of flue-cured tobacco, we adopted ¹⁵N tracing technique and conducted a 2-year experiment from 2014 to 2015 in eastern China. The experiment included three top-pruning points of time: 5th, 25th and 45th day after flowering (DAF), abbreviated, respectively, as TP5, TP25 and TP45. The amounts of plant N derived from fertilizer (Ndff) and soil (Ndfs) were observed during 0–55th DAF. Results showed that top-pruning slowed down the increase of Ndff in tobacco organs, particularly in the leaf, but accelerated the increase of Ndfs dramatically. The proportion of Ndff (%) accounted for the total N reduced dramatically after top-pruning. This reduction might attribute to the selectivity of plant to different N sources as influenced by top-pruning while had little relationship with soil N supply, according to the analysis on the soil total mineral N and mineral ¹⁵N. The average NUE for the 2 years was 32%, 41% and 47%, respectively, for TP5, TP25 and TP45, showing significant (p < 0.05) differences. We concluded that the tobacco preferred to uptake soil N rather than fertilizer N after top-pruning; thus, optimizing the top-pruning time might be one of the approaches to improve the in-season NUE of flue-cured tobacco.     

不同施氮水平对冬小麦季化肥氮去向及土壤氮素平衡的影响

采用田间微区15N示踪技术,研究了冬小麦季化肥氮去向及土壤氮素平衡。结果表明,在供试土壤肥力水平和生产条件下,N 150 kg/hm2的施肥量已达到较高产量,再增加氮肥用量小麦产量不再增加。随着施肥量的增加,地上部吸氮量有所增加,氮肥的表观利用率和农学利用率持续下降,而生理利用率则表现为低—高—低的变化趋势。在低施氮条件下,小麦主要吸收土壤氮的比例高于化肥氮;在高施氮条件下,小麦吸收土壤氮的比例下降。冬小麦收获后,仍有26.7%4~0.6%的氮肥残留在0—100 cm土层中,17.4%2~4.8%的氮肥损失。残留在土壤剖面中的氮肥主要分布在表土层。随着施氮量的增加,土壤氮素总平衡由亏缺转为盈余;土壤根区硝态氮也由播前消耗转为在播前的基础上累加,两个小麦品种表现为相同的趋势。     

基于15N示踪技术的烟田肥料氮素再利用分析

为研究后季烤烟对首季残留氮肥的吸收利用情况,2011年利用蒸渗仪设计了不同灌水量(600、800和1 000 mm)和施氮量(15N双标记NH_4NO_3,90和120 kg/hm~2)试验,并于2012-2014年对烤烟不同器官中15N-N(来源于2011年施入的肥料氮素)累积量、土壤中15N-N含量进行了跟踪观测,同时分析了烤烟对首季肥料氮素再利用率的影响因素。结果表明:1)后季烤烟叶、茎和根中来源于首季的肥料氮素累积量,随首季施氮量的增加而增加,但随首季灌水量增加总体上有所下降;2)后三季烤烟对首季施入肥料氮素的总再利用率为10.79%~14.58%,首季灌水量600 mm、施氮量90 kg/hm~2处理最有利于后季烤烟对其残留肥料氮素的吸收;3)后三季烤烟对首季施入的肥料氮素的平均再利用率,与首季灌水量呈极显著负相关(P0.01),与0~20 cm土壤中首季残留肥料氮素量呈显著正相关(P0.05)。首季灌水主要通过改变其肥料氮素在不同土层中的分配格局,尤其是通过改变0~20 cm土层的肥料氮素含量,来影响后季烤烟对首季施入肥料氮素的再利用,但其具体影响机理仍需进一步明确。综上,后季烤烟能吸收相当比例的前季残留肥料氮素,合理制定前季灌水和施氮制度,对于后季烤烟肥料氮素再利用率的提高至关重要。研究结论可为烟区土壤生态环境的改善及烟草农业的可持续发展提供有益参考。     

A N tracing model to analyse N transformations in old grassland soil

A new ¹⁵N tracing model was developed to analyse nitrogen (N) transformations in old grassland soil. There was a need to develop a new model because existing models such as FLUAZ were not able to simulate the observed N dynamics. The new features of the model are: (a) simulation of heterotrophic nitrification, (b) simulation of dissimilatory nitrate (NO₃⁻) reduction to ammonium (NH₄⁺) (DNRA), (c) release of adsorbed or stored fertiliser N into the available mineral N pools and (d) immobilisation of NH₄⁺ and NO₃⁻ into two separate organic N pools with different re-mineralisation characteristics. The tracing model contains six N pools and nine simultaneous N transformations either at zero- or first-order kinetics. The model is set up in the modelling software ModelMaker which contains non-linear optimisation routines based on the Marquardt-Levenberg algorithm. The model is able to simulate data obtained from triple labelling studies where either the NH₄⁺, the NO₃⁻ or both pools were labelled with ¹⁵N. The flexible modelling environment allows the user to develop the model further.     

Comparison of two chemical methods for extracting residual N fertilizer

Summary Considerable effort has been spent in developing chemical indices to predict N mineralization. However, in spite of numerous studies, the relationship between the index value and plant N uptake has not been as apparent as hoped, and therefore, additional work is required to evaluate the ability of promising new indices to predict the extraction of mineralizable N from soil. The objective of the present study was to evaluate the use of phosphate borate and hot KCl to extract immobilized ¹⁵N-labeled fertilizer, applied 1 and 2 years previously. Soil samples (0–15 cm) were collected on 12 June 1989 from field soil fertilized in either 1987 or 1988 with ¹⁵N-labeled urea. In the laboratory, net N mineralization over 51 days and the amount of N extracted by the phosphate borate and hot KCl methods were determined. In the field, the amount of residual fertilizer and soil plus fixed N in soybeans (Glycine max) at the V5 growth stage were determined on 12 June 1989. The extractability ratio (ER^*) and the mineralizable extractability ratio (MER) were higher for mineralizable N and phosphate borate N for fertilizer applied in 1988 than 1987, while ER^* and MER values for the hot KCl were similar for both application dates. These results suggest that compositional changes occurred which influenced the extractability and mineralization of residual fertilizer applied 1 and 2 years previously, and that the phosphate borate was able to predict these changes while the hot KCl method was not.     

Kinetics of 15N -labelled nitrogen from co-compost made from cattle manure and chemical fertilizer in a paddy field

In order to produce an effective organic fertilizer, cattle manure was cocomposted with chemical fertilizer. And the kinetics of nitrogen uptake by rice plants from the co-compost was investigated using the ¹⁵N labelled co-composts on either cattle manure or chemical fertilizer. As a control, nitrogen kinetics from the mixture of cattle manure and chemical fertilizer without co-composting was investigated. At the early stage, rice growth may have been promoted by co-composting, while, it may have been promoted by the larger N-content of cattle manure at the harvesting stage. The ratios of nitrogen uptake by rice plants and residual nitrogen in soil from the cattle manure and chemical fertilizer were determined by measuring ¹⁵N -atom%. The N -uptake ratios by rice plants from the cattle manure in the co-composted plot were about 2–4 times higher than those from the cattle manure without co-composting. However, the N -uptake ratios from the chemical fertilizer in the co-composted plot were lower than those from the chemical fertilizer without co-composting. The N -content of the rice plants derived from chemical fertilizer without co-composting decreased consistently after 28 d. The nitrogen from chemical fertilizer in the co-compost was absorbed again in the latter period of rice growth. The total nitrogen uptake by rice plants from cattle manure and chemical fertilizer was similar regardless of co-composting. However, co-composting would be advantageous at least· in terms of the following aspects: increase of the N -uptake by rice plants from cattle manure, slow-release ability of nitrogen from chemical fertilizer, decrease of nitrogen loss by denitrification.     

烤烟烟碱合成及其氮素来源与移栽期和氮肥的关系研究

采用田间试验与15N同位素示踪微区试验,在湖北襄樊植烟生态区老湾村(N 31°27,′E 111°14′,海拔1130 m)研究了3个不同移栽期和施用氮肥对烤烟烟碱含量和烟碱氮素来源的影响。研究结果表明,与5月5日移栽相比,推迟移栽期至5月15日2~5日,烟碱含量、烟碱氮占总氮比例以及烟碱肥料氮比例分别平均增加10%8~5%、5%1~10%和21%5~6%;与不施氮相比,施用氮肥提高烟碱含量12%5~9%,提高烟碱氮占总氮比例5%~127%。烤烟烟碱氮占总氮的比例随生育进程逐渐增加,而各部位烟叶烟碱肥料氮比例随生育进程和叶位上升逐渐下降。土壤氮是烟碱氮的主要来源,对烟叶烟碱含量有决定性影响。结果说明在当地条件下,控施氮肥和适当提前移栽期更有利于降低烟叶(尤其是中、上部叶)中的烟碱含量。     

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.     

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.     

Cotton growth and the fate of N fertilizer as affected by saline water irrigation and N fertigation in a drip-irrigated field

The objective of this two-year field experiment was to study the effects of irrigation amount, N rate, and irrigation water salinity on cotton growth and the fate of N fertilizer. The movement of N through the plant-soil system was traced using ¹⁵N-labeled urea. The study consisted of twelve treatments, including two irrigation amounts (405 and 540?mm, I405 and I540, respectively); two N application rates (240 and 360?kg?N/ha, N240 and N360, respectively); and three irrigation water salinity levels [0.35, 4.61 and 8.04?dS/m, representing fresh water (FW), brackish water (BW), and saline water (SW), respectively]. A randomized complete block design was used with three replications. The results showed that cotton biomass, N uptake, and yield increased as irrigation amount and N amount increased; however, all three variables were significantly less in SW than in FW and BW. Plant ¹⁵N recovery rates were greater (i) in the I540 treatments than in the I405 treatments and (ii) in the N360 treatments than in the N240 treatments. Plant ¹⁵N recovery rates in BW were 7.98% and 30.01% greater than those in FW and SW, respectively. Residual soil ¹⁵N increased as N fertilizer amount increased but declined as irrigation amount increased. Residual soil ¹⁵N in BW and SW was 6.02% and 21.44% greater, respectively, than in FW. Total ¹⁵N recovery was significant greater in BW than in FW and SW. The ¹⁵N leaching losses increased significantly with increases in irrigation amount, irrigation water salinity, and N rate. Our study suggests that if appropriate amounts of irrigation water and N fertilizer are used, then brackish irrigation water (4.61?dS/m) will not affect cotton growth, yield and N recovery. In contrast, saline irrigation water (EC?>?8?dS/m) reduces cotton growth, yield, and N use efficiency.     

Fate of residual 15N-labeled fertilizer in dryland farming systems on soils of contrasting fertility

Abstract

Up to 50% of nitrogen (N) fertilizer can remain in soil after crop harvest in dryland farming. Understanding the fate of this residual fertilizer N in soil is important for evaluating its overall use efficiency and environmental effect. Nitrogen-15 (¹⁵N)-labeled urea (165 kg N ha^?1) was applied to winter wheat (Triticum aestivum L.) growing in three different fertilized soils (no fertilizer, No-F; inorganic nitrogen, phosphorus and potassium fertilization, NPK; and manure plus inorganic NPK fertilization, MNPK) from a long-term trial (19 years) on the south of the Loess Plateau, China. The fate of residual fertilizer N in soils over summer fallow and the second winter wheat growing season was examined. The amount of the residual fertilizer N was highest in the No-F soil (116 kg ha^?1), and next was NPK soil (60 kg ha^?1), then the MNPK soil (43 kg ha^?1) after the first winter wheat harvest. The residual fertilizer N in the No-F soil was mainly in mineral form (43% of the residual ¹⁵N), and for the NPK and MNPK soils, it was mainly in organic form. The loss rate of residual ¹⁵N in No-F soil over summer fallow was as high as 48%, and significantly (P < 0.05) higher than that in the NPK soil (22%) and MNPK soil (19%). The residual ¹⁵N use efficiency (RNUE) by the second winter wheat was 13% in the No-F soil, 6% in the NPK soil and 8% in the MNPK soil. These were equivalent to 9.0, 2.0 and 2.2% of applied ¹⁵N. The total ¹⁵N recovery (¹⁵N uptake by crops and residual in 0–100 cm soil layer) in the MNPK and NPK soils (84.5% and 86.6%, respectively) were both significantly higher than that in the No-F soil (59%) after two growing seasons. The ¹⁵N uptake by wheat in two growing seasons was higher in the MNPK soil than in NPK soil. Therefore, we conclude that a high proportion of the residual ¹⁵N was lost during the summer fallow under different land management in dryland farming, and that long-term combined application of manure with inorganic fertilizer could increase the fertilizer N uptake and decrease N loss.     

氮肥施用对作物吸收土壤氮的影响——兼论作物氮肥利用率

在15N标记肥料微区试验中采取了抑制土壤中氮生物固持作用的措施——用长期不施肥试验地、小麦拔节前施15N标记KNO3,成功显示了影响施氮作物吸收土壤氮的另一个过程:肥料氮对土壤氮库的稀释作用(或作物吸收养分过程中肥料氮和土壤氮的库替换作用)的存在;试验中施氮作物较不施氮作物少吸收土壤氮,呈现负的ANI(加入氮交互作用)。生物固持作用和肥料氮对土壤氮库的稀释作用是两个作用相反且交织在一起的过程,其相互抵消、平衡后的最终结果可以是正ANI,也可以是负ANI,决定于两者影响力的强弱。由于受上述过程的影响,用田间试验估测作物氮肥利用率,无论15N标记法或传统的差值法,均不可能获得可信的结果。作为替代方法,比值法可避免上述过程的影响,经本试验检验,结果良好。     

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。     

Effect of long-term fertilization on 15N uptake and retention in soil

We did a pot experiment with three different fertilized soils (no fertilizer (No-F), inorganic fertilizer nitrogen, phosphorus and potassium (NPK), manure plus inorganic fertilizer (MNPK)) from a 19-year fertilizer trial. Three N treatments, (1) no N, (2) 100 mg/kg urea-¹⁵N (N), (3) 50 mg/kg urea-¹⁵N + 50 mg/kg corn straw-N (1/2N + 1/2S), were applied to each soil. The residual soil from the same treatments was used to grow second wheat crop. The MNPK soil had significantly higher nitrogen use efficiency (NUE) in the first growing season, and lower N loss than the NPK, and No-F soils. The 1/2N + 1/2S treatment decreased NUE on each soil, even though the MNPK soil still had highest NUE and lowest N loss. The residual ¹⁵N use efficiency (RNUE) in 1/2N + 1/2S treatment of MNPK soil was higher than NPK and No-F soils. We concluded that long-term application of manure plus inorganic fertilizer increased NUE and decreased N loss.     

Effects of Irrigation Water Quality and Nitrogen Rate on the Recovery of 15N Fertilizer by Sorghum in Field Study

The effects of ¹⁵N-labeled ammonium nitrate on yield, uptake of nitrogen (N) by sorghum (Sorghum sudanense, Piper), and on N remaining in the soil were studied in a field experiment with different N rates (0, 50, and 100 kg N ha^?1) and with two irrigation water qualities, well water (WW) and treated wastewater (TWW). Treated wastewater irrigation increased dry matter and N yield compared to WW. At equal N rates, recovery of ¹⁵N-labeled fertilizer by plants increased with TWW irrigation compared to WW (36% versus 23%). Neither fertilizer rate nor water quality had an effect on the ¹⁵N-labeled fertilizer remaining in the 0- to 60-cm layer of soil. On average 41% in the TWW treatment (49–33%) and 38% in WW treatment was mostly present in the surface 20-cm layer. Losses of ¹⁵N-labeled fertilizer were unaffected by irrigation water quality (35%) and increased with N application rate in TWW (4% versus 31%).     

Translocation and recovery of 15N-labelled N derived from foliar uptake of 15NH3 by rice (Oryza sativa L.) cultivars

Foliar uptake of ¹⁵NH₃ applied at two growth stages (tillering and anthesis) and the subsequent ¹⁵N-labelled vegetative-N distribution in different plant components at maturity was investigated in three rice cultivars, IR-6, NIAB-6 and Bas-385. Rice plants absorbed 22–30% and 18–24% of the ¹⁵NH₃ applied at tillering and anthesis stages, respectively. Of the total ¹⁵NH₃ absorbed at tillering stage, IR-6 and Bas-385 showed higher recovery (71%) in different plant components at maturity as compared to NIAB-6 (48% recovery). At maturity, percent recovery of the ¹⁵NH₃ absorbed at anthesis stage was almost comparable in different cultivars, but it was lower (46–55%) than that absorbed at the tillering stage. Recovery of the absorbed ¹⁵NH₃-N in the soil was negligible and ranged from 0.3–1%. At maturity, the cultivars IR-6 and Bas-385 showed a higher loss (45–53%) of ¹⁵NH₃ absorbed at anthesis than at the tillering stage (29% loss), whereas for NIAB-6, the corresponding figures were comparable for the two growth stages (tillering, 51% loss; anthesis, 49% loss). Results indicated a variable potential of the tested rice cultivars for foliar uptake of atmospheric ¹⁵NH₃ and distribution of ¹⁵N-labelled vegetative-N in different plant components.     

Temporal changes in distribution and composition of N from labeled fertilizer in soil organic matter fractions

Variations in the amount and composition of immobilized nitrogen (N) in major soil organic matter fractions were investigated in a 730-day soil incubation experiment using ¹⁵N-labeled urea and ¹⁵N nuclear magnetic resonance spectroscopy with the cross polarization/magic angle spinning (¹⁵N CPMAS NMR) method. After 730 days, 24.7% of the applied N was recovered from the soil as organic N. The urea-derived N recovered from humic acids and humin decreased from 11.2 and 33.8% of the applied amount after 14 days to 1.6 and 20.4% after 730 days, respectively. When these values were corrected for the microbial biomass (MB) N, they ranged from 9.0 to 1.2% and 28 to 18%, respectively. The proportion of urea-derived N recovered from fulvic acids was low, ranging between 0.4 and 5.8% (with MB N) or 5.6% (without MB N) of the applied amount, whereas that from water-soluble nonhumic substances (WS-NHS; NHS in the fulvic acid fraction) remained high, 28–33% of the applied amount after correction for the contribution of MB N up to day 365, and decreased to 0.9% thereafter. The ¹⁵N CPMAS NMR spectra of humic acids, fulvic acids, and humin showed the largest signal at −254 to −264 ppm, corresponding to peptide/amide N. The proportions of heterocyclic, peptide/amide, guanidine/aniline, and free amino N in the urea-derived humic acid N were 3–7, 83–90, 5–7, and 2–4%, respectively. More than 80% loss of the urea-derived humic acid N did not markedly alter their composition. No time-dependent variations were also observed for the proportions of respective N functional groups in humin N, which were 3–5, 71–78, 12–17, and 6–10% in the same order as above. These results suggest the greater importance of physical stability than structural variation for the initial accumulation of organic N in soil.     

黄壤上烤烟氮素积累、分配及利用的研究

田间条件下,利用同位素15N示踪技术于黄壤有机质含量分别为19.2和40.7 g/kg和当地推荐最佳氮肥用量基础上,设15N用量分别为105和82.5 kg/hm2的情况下,研究了两个试验点烤烟15N积累、吸收比例、氮素利用率及15N在各器官分配。结果表明,在二种有机质含量的黄壤上,烤烟15N吸收规律相似,于烤烟移栽后3~5周内,烟株吸收15N较少,5周后15N积累量明显增加,到移栽后13周达到高峰,肥料15N吸收时间拖后;二种土壤上,肥料15N在整个生育期内积累量分别为28.41和26.55 kg/hm2。烟株于移栽后3~5周来自肥料15N占吸收总氮的比例为53.84%～71.33%,氮（15N）肥利用率为1.11%～7.34%;到烟叶采收结束（移栽后17周）时,烟株来自肥料15N占吸收总氮的比例为28.69%～29.75%,氮（15N）肥利用率为27.06%～32.18%。各个部位烟叶采收结束时,二种土壤上,肥料15N在上部、中部、下部烟叶及茎和花积累分别占吸收肥料总15N的35.08%~35.26%、25.87%~26.19%、17.92%~18.25%和22.73%~24.49%,肥料15N主要集中在中、上部烟叶。可见,肥料氮吸收时期拖后,土壤后期供氮能力强和中上部烟叶肥料氮比例较高是黄壤烟区烤烟氮素营养存在的主要问题。     

Incorporation of 15N into various nitrogenous compounds in intact soybean nodules after exposure to 15N2 gas

The intact nodules attached to the upper part of soybean roots were exposed to ¹⁵N₂ and the incorporation of ¹⁵N into various soluble nitrogen constituents was investigated. Results indicated that ammonia, a primary product of N₂ fixation, was located in more than two compartments. Ammonia reduced from N₂ gas seemed to be incorporated firstly into glutamine especially amido-group nitrogen. Newly fixed nitrogen was secondly incorporated into glutamic acid and alanine in this sequence. These results suggested that fixed ammonia was assimilated by glutamine synthetase/glutamate synthase pathway. Turn-over rate of allantoin plus allantoic acid and serine was relatively high, although apparently these compounds were not primary products of newly fixed ammonia. ¹⁵N content of allantoin was always higher than that of allantoic acid. ¹⁵N incorporation to aspartic acid and asparagine was relatively slow, especially in early period. In bacteroid fraction there is much amount of ammonia comparing with other compounds, while allantoin and asparagine were presented exclusively in cytosol. ¹⁵N was incorporated into nitrate within a few minutes especially in bacteroids.     

Quantitative analysis of the initial transport of fixed nitrogen in nodulated soybean plants using 15N as a tracer

The quantitative analysis of the initial transport of fixed isotope 15-nitrogen (¹⁵N) in intact nodulated soybean plants (Glycine max [L.] Merr. cv. Williams) was investigated at the vegetative stage (36 days after planting, DAP) and pod-filling stage (91 DAP) by the ¹⁵N pulse-chase experiment. The nodulated roots were exposed to N₂ gas labeled with a stable isotope ¹⁵N for 1 h, followed by 0, 1, 3 and 7 h of exposure with normal air. Plant roots and shoots were separated into three sections (basal, middle and distal parts) with the same length of the main stem or primary root. Approximately 80 and 92% of fixed N was distributed in the basal part of the nodulated roots at the vegetative and pod-filling stages by the end of 1 h of ¹⁵N₂ exposure, respectively. In addition, about 90% of fixed ¹⁵N was retained in the nodules and 10% was exported to root and shoot after 1 h of ¹⁵N₂ exposure at 91 DAP. The percentage distribution of ¹⁵N in the nodules at the pod-filling stage decreased from 90% to 7% during the 7 h of the chase period, and increased in the roots (14%), stems (54%), leaves (12%), pods (10%) and seeds (4%). The ¹⁵N distribution was negligible in the distal root segment, suggesting that N fixation activity was negligible and recycling fixed N from the shoot to the roots was very low in the initially short time of the experiment.