Mr. Aritra Saha and

Dr. Pabitra Kr. Mani

DEPARTMENT OF AGRICULTURAL CHEMISTRY AND SOIL SCIENCE

FACULTY OF AGRICULTUREBIDHAN CHANDRA KRISHI VISWAVIDYALAYA

Background information

Industrialized country : Net positive nutrient balance and soil fertility

build up

Tropical countries : Net mining of nutrient

(Sanchez, 1994)

Contd..

Isotope ??

Atoms with the same number of protons and electrons but differing numbers of

neutrons. e.g. 12C6

Stable Istopes Radioactive isotopes

Stability no. of neutrons(N) is quite similar to no. of protons(z) or N/Z<1.5

Relative abundance of 15N

Element Isotope Abundance %

Relative abundance

%

International Standard

Absolute abundance

of the standard(Rstandard)

Nitrogen 14N 99.635 7.1 Atm. N(air) 15N : 14N = 0.0036765

15N 0.365

Source: E.W. Sulzman, 2007

Measurement of isotopes

Diagrammatic representation of CF and Dual inlet IRMS.

(Dawson & Brooks, 2001)

IRMS

Sample preparation

• At present day – Combustion method is used to convert total sample N to N2 gas.

• Earlier it was done in 2 steps - Sample N converted to Ammonium by

Kjeldahl process - Ammonium is converted to N2 by alkaline

Na-hypobromite (Hauck and Bremner, 1976)

Notation & terminology

R = ratio of heavy-to-light (typically, rare-to abundant) isotope,

Rsample = ratio in the sample

Rstandard = ratio in the standard

As tracers isotopes are at elevated level, so it expressed as:

Notation & terminology

ᵟsa/istd = isotopic value of the sample relative to the internationalstandard, ᵟsa/ws = isotopic value of the sample relative to the working standard, ᵟws/istd = the isotopic value of the working standard relative to the international standard (all values in ‰; Craig 1957)

•+ve delta value the sample has more heavy isotope than the standard•-ve delta value the sample has less of the heavy isotope than the standard.

Stable Isotope based techniques

• Natural abundance studies- preferential biological, chemical & physical exploitation of light.(e.g. 14N, 16O) over heavy one( 15N, 18O)

• Enrichment techniques – Tracing 15N flux of various N species – estimation of the relative contribution of various processes to the production of the N species in interest.

(Hauck and Bremner, 1976)

Advantages of using 15N According to Hauck and Bremner (1976),

1. Obvious advantage is inherent non-radioactivity.

2. Stability in environment is not limited by time i.e. there is no isotope decay with time and can be used for long time experiment.

3. Their use doesn’t pose any health hazards or any disposal problem on biological system.

4. No permission is needed to carry out experiment using 15N tracer technique without any licensing and radioactivity monitoring.

5. Less need of background information of the experimental plot where the tracer will be introduced.

Use of 15N in soil-plant nutrition study

• Tracing studies: Nitrogen cycling, Nitrate pollution in groundwater.

• Fractionation studies: N2 fixation

How much of the fertilizer is uptaken? How much is lost to the environment?

Use of 15N in soil-plant nutrition study Nitrogen use efficiency

(i)recovery of fertilizer N, either in grain (RENG) or in total above-ground biomass (RENT) in the current crop.

(ii) a part of the applied N that is left behind (immobilized) in the soil and becomes available to subsequent crops.

Making a final judgment on REN, both the components should be considered. (Ladha et al.,2005)

Use of 15N in soil-plant nutrition study

• 15N labelling also reveals the differences in the uptake of soil N between fertilized with N and those unfertilized.

Fertilized plant take up more N due to priming effect or added nitrogen interaction(ANI)

• Direct and accurate evaluation of the fertilizer contribution to the crops.

• Fate of non-efficient fraction

The basic Ndff approach

•%Ndff = (% N excess in sample/%N excess in fertilizer)*100•% Ndfs = % of N in the plant tissue from soil

= (100-% Ndff)

(Dourado et al., 2009)

Scenarios that can arise in field experiments using 15N labelling. Witty, 1983

Field experiment showing (a) chickpea plants growing within a steel microplot for 15N labelling and outside the microplot (unlabelled), (b) soil sampling from within a faba bean microplot and (c) deeper soil coring from within the faba bean microplot (IAEA Tech Bull, 2003)

Case studies using 15NTable 2: Percent N, % N atom excess, % Ndff of plant parts of

corn under Till and no-till(NT)

(Ismaili et al. 2015)

Means in columns with different a, b, c letters are significantly different at p = 0.05.

Table 3: Percent N, %15N,%Ndff at harvest time of corn and different soil depths.

(Ismaili et al. 2015)

Table 4: Effect of timing of urea-N application on N derived from fertilizer (Ndff), N derived from soil (Ndfs) and Added Nitrogen Interaction (ANI) (Priming effect).

(Bronson et al.,2000)

† ANI = Ndfs in fertilized pots—Ndfs in control pots; significantly different from zero at P < 0.01 in all cases.

‡ ANI mg15N taken up in 14N-fertilized pots—mg 15N taken up in control plots; significantly different zero P< 0.01 in all cases

Table 5: Effect of timing of N application on nitrogen content of rice grain and straw and atom % 15N of rice grain, straw and soil organic matter at physiological maturity.)

(Bronson et al.,2000

† Plus fine roots.‡ Least significant difference at the 0.05 probability level.

Table 6: N15 balance in rice and soil at physiological maturity as affected by timing of application.

(Bronson et al.,2000)

† Plus fine roots.‡ Least significant difference at the 0.05 probability level.

Case studies using 15N• Application effects of sewage sludge (SS) on growth indices,

yield, and nutrient uptake in Komatsuna (Brassica campestris var. perviridis) grown in a low fertility soil were investigated and compared with those of chemical fertilizer (CF) and no-fertilizer (NF) treatments. The N-use efficiencies of CF and SS were 19.7% and 12.1%, respectively, of the applied N. Therefore, the relative efficiency of the sewage sludge to chemical fertilizer was 61.5%. (Asagi et al. 2008)

Case studies using 15N• Ghoneim (2008) reviewed nitrogen dynamics and fertilizer use

efficiency in rice using the N15 isotope techniques in Egypt and observed that fertilizer use efficiency calculated by the 15N dilution method tended to be higher for CF (Chemical Fertilizer) than DAOR (digested anaerobic organic residues).

• A significant amount of N fertilizer (average: 30%) was apparently lost from the soil-plant system through ammonia (NH3) volatilization. Following DAOR application, the pH increased by 1.0 to 1.2 units in the top 5 cm of the soil, resulting in high NH3 volatilization in the first 2 days of the experiment. The NH3 volatilization accounted for the decrease in soil ammonium-N (NH4-N) content.

Case studies using 15N

• Widory et al. (2005) tracked the sources of nitrate pollution in groundwater pollution through mineral fertilizers, wastewater and animal manure.

• Jagadeeswaran et al. (2004) studied on NUE in turmeric at coimbatore and observed recovery of N in turmeric was in favour of 4 splits at 180 days growth stage (19.46%) as well as harvest (30.76%).

Case studies using 15N• Ko-H-J (2003) investigated that the nitrate

contamination sources in different agricultural system in Korea Republic, by using nitrogen isotope ratios.

• The nitrate concentrations of groundwater in livestock farming area were higher than those in conventional and organic farming area and exceeded the national drinking water standard of 10 mg N/litre. The delta 15N ranges of chemical fertilizer and animal manure were -3.7~+2.3 per mil and +12.5~26.7 per mil, respectively.

Case studies using 15N

• Bronson and Fillery (1998) conducted a study on fate of nitrogen-15-labelled urea applied to wheat on a waterlogged texture-contrast soil, recorded that denitrification measured by 15N-chamber methods, was the largest loss mechanism identified during waterlogging.

Recovery of 15NO3-N in texture-contrast soil profile a) pre-waterlogging, and b) after water logging at 28 d after fertilization (DAF)

Increasing the period between urea fertilization and water logging from 6 – 12 d resulted in NO3-N levels in the 0-5 cm soil layer increasing from 9 – 19% of the applied 15N

Prior to waterlogging, nearlyall inorganic N remained in the surface 10 cm. By 28 DAF leaching of NO3 after water logging at 28 DAF was substantial Bronson and Fillery (1998)

•A study on effect of green manuring in reducing the gaseous losses of applied fertilizer nitrogen (in form of 15N) from rainfed rice soil by revealed that there was about 42 % loss of fertilizer nitrogen applied to alluvial rainfed rice soil of West Bengal.

Bhattacharyya and Mandal (1997) Works at BCKV

Constraints • High cost and unavailability of 15N labelled fertilizer.

• Difficulty in N isotope ratio analysis. It requires highly skilled technicians and operational procedures.

• Ndff equation doesn’t give the route taken by the nitrogen in going from pool A to pool B.

• The 4th reservation concerns the amount of information Ndff eq. yields on the fate of unlabelled pool in the soil, the so called soil N pool. In fact it doesn’t gives any info about soil N. It is possible to recovery of soil N in the plant by subtracting that recovered from the added label from the total N content of the plant.

Future scope of research

•Develop methods for distinguishing between N in the various soil N pools.•Define and quantify reaction mechanisms whereby N becomes stabilized.•Establish the relationship between mineralization-immobilization turn over and gaseous loss of soil and fertilizer N.•Quantify the relationship between soil organic and inorganic N pools as influenced by soil type, cropping system, climate, residue management practices.•Determine the long time fate of immobilized N under different crop management practices.

Pictures of the installation of the Lysimeter

The Agro-Lysimeter visualizes the availability of water and fertilizer, and measures evapotranspiration and leachate.

Conclusion