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NT2605 Final Report WP3 Optimum use of Agrotain________________________________________________________________________________

Component report for Defra Project NT2605 (CSA 6579)

WP3 Optimum use of nBTPT (Agrotain) urease inhibitor

Lead Authors

Dr Catherine J Watson and Dr Nasir A Akhonzada, Queen’s University of Belfast

November 2005

NT2605 Final Report Optimum use of Agrotain.doc 1


Contents

1. EXECUTIVE SUMMARY………………………………………………………………………………….4

2. INTRODUCTION…………………………………………………………………………………………...5

3. EXPERIMENTAL DESIGN, TREATMENTS AND METHODS………………………………...…..…8

4. RESULTS AND DISCUSSION………………………………………………………………………….12

5. CONCLUSIONS……………………………………………….………………………………………….32

6. FUTURE RESEARCH REQUIREMENTS………………………………………………………...…...34

7. REFERENCES……………………………………………………………………………………………35

8. APPENDICIES…………………………………………………………………………………………….37



Abbreviations

nBTPT N-(n-butyl)-thiophosphoric triamide urease inhibitorNBPTO N-(n-butyl) phosphoric triamideNMP N-methyl-pyrrolidoneHPLC High Pressure Liquid ChromatographyCEC Cation exchange capacityN NitrogenNH3 AmmoniaNH4 AmmoniumNO3 NitrateNO2 NitriteQUB Queen’s University, BelfastUAN Urea ammonium nitrate solutionppm Parts per millionw/w weight by weightKCl Potassium chlorideMAFF Ministry of Agriculture, Fisheries and FoodTRAACS Technicon Random Access Automated Chemistry SystemLC Liquid chromatographyMS Mass Spectroscopy



1. Executive Summary

The current study evaluated the effect of rate of application of the urease inhibitor nBTPT, trade name ‘Agrotain’, (0, 100, 250, 500, 750 or 1000 ppm w/w active ingredient nBTPT) on NH3 volatilisation in three formulations of urea (coated, added to the urea melt or in UAN solutions), at three temperatures (5, 15 and 25C) and with four contrasting soil types. Ammonia volatilisation was studied for up to 21 days after surface N application, using ventilated soil enclosures, under laboratory conditions. Asymmetrical sigmoidal curves were fitted to cumulative daily NH3 volatilisation data and the time of maximum rate of loss (Tmax) and maximum daily loss rate were estimated. In addition, the stability of nBTPT in different formulations was investigated by high pressure liquid chromatography.

Agrotain was highly effective in lowering NH3 volatilisation from urea. The average % inhibition over all soils, temperatures and formulations was 61.2%, 69.9%, 74.2%, 79.2% and 79.8% for the 100, 250, 500, 750 and 1000 ppm nBTPT concentration, respectively. The % inhibition with nBTPT was lower at 15C compared to 5C or 25C and was lower in UAN solution than in granular products. It was suggested that this effect may be due to the speed of formation of the oxygen analogue in soil and/or its stability. There was little difference between the melted and coated granular products in reducing NH3 loss or in soil N transformations. There was little additional benefit in using concentrations of nBTPT above 500 ppm in any formulation. The inhibitor not only lowered total NH3 volatilisation and the maximum daily rate, but delayed the time of maximum rate of loss (Tmax). Under field conditions, delaying Tmax would increase the opportunity of rain falling to move urea below the soil surface and lower NH3 loss.

Ammonia loss from unamended urea varied with soil type and temperature and ranged from 8.2% to 31.9% of the N applied. It was not possible to explain the variation in NH3 emission with temperature or soil properties.

The stability of nBTPT was highly dependent on temperature in all fertiliser formulations, with the rate of degradation being greatest at 25C. nBTPT was much less stable in the coated urea fertilisers than in either the melted or UAN products. The nBTPT in the melted products maintained in bulk quantities at 4C was stable at all concentrations.



2. Introduction

2.1. The NT26 Research programme

The NT26 research programme was set up by Defra to investigate the nitrogen (N) loss pathways, the environmental and economic impacts, and the response of agricultural and horticultural crops to different forms of fertiliser-N. The NT2605 project was part of a suite of projects in this programme as shown below (Final report submission dates shown in brackets).

NT2601 Desk study reports on: Nitrogen fertilising materials (June 2003) Production and use of nitrogen fertilisers (August 2003)

NT2602 Desk study report on: Evaluation of urea-based nitrogen fertilisers (October 2003)

NT2603 Report of field studies (2002/03 cropping season): The behaviour of some different fertiliser-N materials (March 2004)

NT2604NT2606

Facilities construction: Ammonia emissions from nitrogen fertilisers – wind tunnel construction

(March 2004)

NT2605 This project

NT2610 Report of field studies (led by Silsoe Research Institute): Spreading accuracy of solid urea fertilisers (August 2005)

The following leading UK agri-environment research organisations participated in all NT26 projects (except NT2610), including the NT2605 project reported here.

ADAS UK Ltd Edinburgh University (EU) Warwick HRI (HRI) Institute of Grassland and Environmental Research (IGER), North Wyke Queens University, Belfast (QuB) Rothamsted Research (RR) SAC Commercial Ltd (SAC)

The project was led by Peter Dampney, Principal Research Scientist, ADAS Boxworth Research Centre, Cambridge who was the main point of contact with the Defra NT26 Steering Group.



2.2. The NT2605 project

The NT2601, NT2602 and NT2603 projects provided the basis for the field experimental and other work carried out in NT2605 in cropping seasons 2003/04 and 2004/05. The overall aim of the project was to develop working decision support systems (DSS) to evaluate the agronomic, environmental and economic impacts that would result from changes in the use of different fertiliser-N materials in UK agriculture. More specifically, project work packages (WP) covered the following topic areas:-

WP1a To investigate crop responses to different fertiliser N forms.

WP1b To generate robust ammonia emission algorithms and emission factors for predicting the loss of ammonia following application of different fertiliser N forms under a range of crop, soil and environmental conditions. To evaluate the relationship between ammonia loss and crop N use efficiency as a potential basis for revising current national standard nitrogen fertiliser recommendations (Defra, 2000).

WP2 To generate robust nitrous oxide emission factors for predicting losses following application of different fertiliser N forms under contrasting crop, soil and environmental conditions.

WP3 To determine the optimum formulation method, addition rate and method of use of urea treated with the urease inhibitor nBTPT (Agrotain), to maximise its ammonia abatement potential and efficiency of N use by crops, whilst minimising any adverse phytotoxic effects.

WP4 To assess the risk of ammonium-N, nitrite-N or urea-N losses to surface waters and groundwaters following the application of urea-based N fertilisers.

WP5 To assess the potential for urea or urea+Agrotain to cause phytotoxic effects during establishment, in growing crops, or in marketable produce.

WP6 To construct a decision support system that will assess the economic impacts of changes in the availability of different forms of N fertiliser on different farm types and UK agriculture.

WP7 To estimate and evaluate the agronomic, environmental and economic impacts at both farm and national levels that would result following different hypothetical scenarios concerning the availability of N-containing fertilisers to UK farmers.

Reporting of the NT2605 has been structured into a suite of 8 component reports, one for each work package plus an over-arching Executive Summary for the whole project. Each report is self contained with its own Executive Summary, but interacts with data and conclusions from other WPs where appropriate.



2.3. WP3 Optimum use of nBTPT (Agrotain) urease inhibitor

Fertiliser urea can be an inefficient N source due to rapid hydrolysis by the soil enzyme urease, leading to NH3 volatilisation. The efficiency of urea can be improved by soil incorporation or by use of urease inhibitors. As 97% of all N fertilisers used in the UK are top-dressed to crops, there is little scope for the widespread use of soil incorporation as a mitigation strategy to lower NH3 emissions from urea. Urease inhibitors delay the rate of urea hydrolysis and hence prevent localised zones of high pH, which are conducive to NH3

volatilisation. Many compounds have been evaluated as urease inhibitors (Martens and Bremner, 1984, Mulvaney and Bremner, 1978, Watson, 2000), however, few meet the requirements of being effective at low concentrations, non-toxic, stable, inexpensive and compatible with urea. N-(n-butyl) thiophosphoric triamide (nBTPT), a structural analogue of urea, is currently the most promising. Its urease inhibitory activity in soil is associated with the activity of its derivative, the oxygen analogue, N-(n-butyl) phosphoric triamide (NBPTO). The factors that affect the rate of conversion to the oxygen analogue have not been fully elucidated and will probably depend on a number of biotic and abiotic soil properties. The conversion to nBTPT to its oxygen analogue is generally rapid, occurring within minutes or hours in aerobic soils (Byrnes and Freney, 1995).

Until recently, the only European work with nBTPT had been on grassland by Watson (2000) in Northern Ireland, who showed that nBTPT had considerable potential for improving the efficiency of urea under temperate conditions. Coating nBTPT onto urea granules lowered NH3 volatilisation and increased N recovery and dry-matter yield of ryegrass compared with urea alone. The inhibitor was active at low concentrations and there appeared to be little benefit in using concentrations above 0.1% (w/w) nBTPT (Watson et al., 1994). The beneficial effects of coating urea with nBTPT have been confirmed in recent field trials with grassland and tillage land during 2003, in the Defra funded project NT2603 (Dampney et al., 2004).

In considering the potential for use of nBTPT in Europe, further work is required to evaluate alternative formulation methods and to determine the optimum rates to reduce NH3 loss. The inhibitor can be used to coat urea granules, be added to the urea melt during manufacture, or be added to urea ammonium-nitrate (UAN) solutions prior to surface application to soil. There is no published information on the efficacy of nBTPT to lower NH3 loss when added to the urea melt or used in UAN solutions. The stability of nBTPT in different formulations under different storage conditions is also an important consideration requiring further investigation.

The current study aimed to evaluate the effect of rate of nBTPT (0, 100, 250, 500, 750 or 1000 ppm w/w) on NH3 volatilisation in three formulations of urea (coated, added to the urea melt or in UAN solutions), at three temperatures (5, 15 and 25C) and with four contrasting soils. Ammonia volatilisation was studied for up to 21 days after N fertiliser application, using ventilated soil enclosures, under laboratory conditions. The stability of nBTPT in different formulations was investigated by high pressure liquid chromatography.



3. Experimental design, treatments and methods

3.1. Soils

Representative surface samples (0-10 cm) of two arable (ADAS Gleadthorpe, GL and Boxworth, BX) and two grassland soils (ADAS High Mowthorpe, HM and IGER North Wyke - Debathe site, DB) with different chemical and physical properties, were collected in February 2004. These soils were used in Work package 1b, thus providing a comparison of field and laboratory based measurements of ammonia emissions. The field moist soils were air dried at 30ºC for a minimum of 18 hours and coarsely sieved through a 6 mm sieve to remove large stones and plant debris. A 0.5 kg sub-sample of each soil was ground (2 mm) prior to other soil analyses.

3.2. Soil analyses

Soil chemical (pH, cation exchange capacity, loss on ignition, % C, extractable P, K and Mg) and physical analyses (% sand, % silt and % clay) were determined according to standard methodology (MAFF, 1986) and are shown in Table 1. Soil extractable P, K, and Mg were expressed on a soil volume basis (i.e. mg l -1). The gravimetric moisture content of the soils at field capacity was determined using the Haines method (Rowell, 1994), as the retained water capacity under a tension of 0.05 bar. Urease activity was measured on soil rewet to field capacity and allowed to equilibrate for one week at 15C. Urease activity was based on the determination of NH4

+-N released during soil incubation with THAM (Tris (hydroxyl-methyl) aminomethane) buffer and urea solution at 37ºC for 2 hours (Tabatabai, 1982). The NH4

+-N released was determined using an automated continuous flow wet chemistry analyser (Skalar, SAN++), following soil extraction with 2M KCl containing 5 μg ml -1 PMA (phenyl mercuric acetate) to stop urease activity.

3.3. Fertiliser

The granular products were produced with Agrotain added to the urea melt, by fluidized bed granulation, to give target concentrations of 0, 100, 250, 500, 750 and 1000 ppm nBTPT on a urea weight basis. Agrotain refers to the clear green liquid containing 25% nBTPT as the active ingredient. The nBTPT was in a mixed solvent consisting of 10% by weight of N-methyl-pyrrolidone (NMP) with the balance consisting of propylene glycol. No Agrotain was added to the 0 ppm nBTPT product so there was no NMP or propylene glycol present in these granules. The melted products were manufactured on 22nd January 2004 and analysed for nBTPT concentration on 28th January 2004. Approximately 2 kg of each of the granular products was delivered to Queen’s University, Belfast (QUB) on 6 th February 2004 and was stored in a cold room at 4C in sealed bags, prior to use. The same batch of product was used throughout work package 3.

Urea was spray impregnated (i.e. coated) with Agrotain at ADAS Boxworth, to give target concentrations of 0, 100, 250, 500, 750 and 1000 ppm nBTPT on a urea weight basis. To achieve the different concentrations, a standard volume (24 mls) of Agrotain or Agrotain diluted with propylene glycol was used to spray a 6 kg batch of urea using a high velocity centrifugal seed dressing machine. The coating of the urea products commenced on 13 February 2004. However, as there was concern that the coated products may not be stable



over the long-term, additional products were coated on 2 April 2004, 21 May 2004, 30 July 2004 and 11 January 2005. The coated products were stored in a cold room at 4 C in sealed bags, prior to use.

The urea ammonium nitrate (UAN) solution was sourced by ADAS (Nuram 37 product, 37 % N w/v, bulk density 1.289 kg l-1) and Agrotain was added immediately prior to use to give target concentrations of 0, 100, 250, 500, 750 and 1000 ppm nBTPT on a urea weight basis.

The concentration of nBTPT in each of the fertiliser products was validated by high pressure liquid chromatography (HPLC), with ultra violet (UV) detection, at QUB at the start of each experimental run (Appendix 1). Independent analyses of nBTPT showed good agreement with QUB. There was also good agreement between nBTPT concentrations determined by LC-UV and LC-MS (liquid chromatography-mass spectroscopy).

In February 2004 the variability of nBTPT concentration in the granular products (melted and coated) was investigated by analysing 10 x 100 mg samples (to 10 mls water) of each product (melted and coated) and target concentrations (0, 100, 250, 750 and 1000 ppm nBTPT). Results are shown in Appendix 2. The nBTPT concentration in the coated products was more variable that in the melted products, as indicated by the higher standard deviation. In addition, analytical reproducibility was investigated on selected products by analysing the same solution ten times, and was found to be excellent (Appendix 3).

3.4. Experimental design

Each NH3 volatilisation run consisted of four soils (BX, GL, HM & DB), six concentrations of the inhibitor nBTPT (0, 100, 250, 500, 750, 1000 ppm) and three formulations (Agrotain impregnated onto urea granules (coated), Agrotain added to the urea melt (melted) and Agrotain added to UAN solution). In addition, two unfertilised samples of each soil were also incubated in each run to act as controls. Each experimental run required 80 jars of soil, which were arranged in a completely randomised design over five shelves in the cabinet; with 40 jars being ventilated from the right hand side and 40 jars being ventilated from the left hand side of the cabinet. Using three temperature regimes (5ºC, 15ºC and 25ºC) and four replicates required twelve separate runs each lasting up to 21 days. An error in the volume of UAN applied resulted in 2 runs being repeated and the number of replicates for the UAN treatment being reduced from 4 to 3. Statistical analysis showed that there was no significant positional effect within the cabinet on any of the parameters studied.

3.5. Measurement of NH3 volatilisation

Ammonia volatilisation was studied for 1521 days after fertiliser application in a dark controlled environment cabinet using ventilated enclosures at sufficient flow rates to promote maximum NH3 removal (11 changes of headspace volume per minute). Air-dried soil was placed in cylindrical screw-top plastic jars (80 mm diameter, 85 mm height) and re-wet with deionised water to field capacity. The weight of each soil type per jar was different, which ensured a standard headspace volume of 176 cm3 in each jar. The moist soil was allowed to equilibrate in loosely capped jars at 15ºC for one week before the fertiliser was applied. The fertiliser granules were 2.8 – 3.35 mm in size and were uniformly applied to the soil surface to give 50 mg N per treatment (equivalent to approximately 100 kg N ha -1). The UAN



solutions containing the required concentration of nBTPT were mixed immediately prior to application to the soil surface using a pipette, to give 50 mg N per jar.

The lid of each jar was fitted with an input and an output port. Air at a high relative humidity (60- 65%) was blown over the soil surface in each jar using an air compressor and flow meters, at a rate of 2 l minute-1 into traps containing 40 mls of 20 mM orthophosphoric acid, which were replaced daily. The acid was transferred to labelled screw top 60 ml plastic jars and the weight recorded. The samples were stored at 4ºC, prior to analysis of NH3-N using a TRAACS 800 continuous flow analyser (Bran and Luebbe, 1989). The total NH3 loss from the soils receiving no fertiliser was very low and averaged 0.006 mg N over 21 days. The NH3-N volatilised was expressed as a percentage of that applied, after adjusting for the controls.

Each jar was weighed prior to the application of fertiliser and at the end of the incubation to determine the moisture loss (by weight difference). At the end of the incubation the total soil in the jar was extracted in 2M KCl and NH4

+-N and NO3- -N concentrations in the soil extracts

were determined using an automated continuous flow wet chemistry analyser (Skalar, SAN++

2003). Any NO2--N present was included in the NO3

--N determinations. Residual urea was determined colorimetrically (Mulvaney and Bremner, 1979). The KCl soil extracts were stored at 4ºC and analysed within one week. All N concentrations were expressed as a percentage of that applied, after adjusting for the mineral N concentrations in the control samples.

3.6. Stability of nBTPT during storage

The stability of 1000 ppm nBTPT was investigated during 2004 in the coated, melted and UAN products stored in a cool dry fertiliser store. The melted product was the same as that used in the experimental runs to measure NH3 volatilisation. It was manufactured on 22 January 2004. The coated product was spray impregnated with Agrotain by ADAS on 13 February 2004. Both granular products were stored in bulk in sealed plastic bags at 4C until 24 March 2004.

Sub-samples (2 x 350 g) of the coated and melted products at 1000 ppm nBTPT were placed into a cool dry fertiliser store in screw top plastic jars on 24 March 2004. The fertiliser store was unheated and the temperature fluctuated according to the ambient outside air temperature. Duplicate 5 g samples were taken at approximately weekly intervals for the first two months and then every 3-6 weeks thereafter. The last sample was taken on 26 August 2005. Daily mean, maximum and minimum temperatures were recorded in the fertiliser store and are shown in Appendix 4.

Ten litres of a UAN solution containing 1000 ppm nBTPT was freshly prepared on 24 March 2004 and stored in a sealed plastic container. A 100 ml sub-sample was analysed in duplicate at the same time as the granular products. In addition, sub-samples of the coated and melted products used in WP3 were maintained at 4C and analysed for nBTPT, at the same time.

To investigate the temperature effect a further storage trial was established during 2005 at 4C, 15C and 25C. Samples were also stored in the fertiliser store for comparative purposes. Urea was coated with 500 ppm nBTPT on 11 January 2005, by ADAS. On 21



March 2005, 200 g samples were placed in sealed plastic bags at 4 C, 15 C, fertiliser store and 25 C. A further sample of the coated material was vacuum packed in small packages (30 g each) and placed at the 4 different temperature regimes, to study the influence of moisture and /or air, on decomposition. Melted product at 682 ppm nBTPT was manufactured on 22 January 2004 (at the same time as the other WP3 samples) and stored at 4C before being re-packaged and placed in 200 g sealed plastic bags at the 4 temperatures on 21 March 2005. This was called the ‘Old melted’ product.

In addition, new melted product at 500 ppm nBTPT was manufactured in April 2004, by using an improved fluidised bed granulation procedure. Further 200g sealed plastic bags of this product were placed at the 4 different temperatures on 21 March 2005. This product was called ‘New melted’ as opposed to the ‘Old melted’ product, which had been stored at 4 C for over a year. One litre solutions of UAN at 500 ppm nBTPT were prepared on 21 March 2005 and placed at each of the temperatures. Duplicate 5 g sub-samples of each of the granular products and 20 mls of UAN were taken at 3 week intervals for nBTPT analysis, from March to August 2005.

3.7. Statistical analysis

The data was analysed using an analysis of variance with temperature, soils, rate of nBTPT and formulation as factors, having removed control values. Observed NH3 loss was expressed as a % of the N applied.

Asymmetrical sigmoidal curves were fitted to cumulative daily NH3 volatilisation data for each of the four replicates per treatment and for each soil. The time of the maximum rate of loss (Tmax) and maximum daily loss was estimated from the sigmoidal curve and the factors compared. The residual maximum likelihood (REML) procedure was used, because when fitting the sigmoidal curves several observations were excluded due to poor fit estimates and therefore the factor levels were unbalanced.

The % inhibition of NH3 loss by nBTPT over the incubation was calculated for each jar, at each inhibitor concentration, from [(C-T)/C] x 100, where C= NH3 loss from urea alone and T= NH3 loss from urea amended with the inhibitor. The total NH3 loss data and the mineral N fractions (including urea) remaining in the soil at the end of the incubation were evaluated using an analysis of variance and statistical significance was evaluated by an F-test. Paired comparisons were evaluated to determine differences in the means for each factor using least significant differences (LSD’s).



4. Results and discussionThe four soils used in this study showed a wide range of chemical and physical properties (Table 1). For example, cation-exchange capacity ranged from 86 to 358 meq kg -1, and sand content ranged from 21.6% to 82.8%. During the incubation the soils dried out. Soil moisture loss was the % change in soil weight. Information on the rate of soil moisture loss was not available, only the total % loss during the incubation. The % soil moisture loss was significantly higher at 25C than at 5C (Table 2), and with the GL and DB soils compared to HM and BX. This is related to the high % sand content of the GL (82.8%) and DB (60.3%) soils.

4.1. Ammonia loss

Total NH3-N loss for each soil, formulation and rate of nBTPT at 5C, 15C and 25C is shown in Figures 1, 2 and 3, respectively. Total NH3-N loss from the unamended granular urea varied with soil type and temperature and ranged from 8.2% of the N applied for the BX soil at 5C (Fig. 1b) to 31.9% for the DB soil at 15C (Fig. 2b). This is within the range of NH3

emissions measured from urea in the NT26 field studies; mean emission factor of 27% from grassland (range 10-58%) and 22% from tillage land (range 2-43%).

The effect of nBTPT concentrations on daily NH3 loss for each soil, at each temperature and for each formulation is shown in Appendix 5. nBTPT was highly effective in lowering NH3

loss, even at the lowest concentration of 100 ppm. The average % inhibition over all soils, temperatures and formulations was 61.2, 69.9, 74.2, 79.2 and 79.8% for the 100, 250, 500, 750 and 1000 ppm nBTPT concentration, respectively. There was little further reduction in NH3 loss with concentrations above 250 ppm nBTPT. The inhibitor not only lowered total NH 3

volatilisation and the maximum daily emission rate, but delayed the time of maximum rate of loss (Tmax) (Table 3). Under field conditions, delaying Tmax would increase the opportunity for the fertiliser to interact with the soil (i.e. by rainfall washing the fertiliser into the soil), reducing NH3 loss. The average % inhibition measured here is in good agreement with results from the 31 field measurements on grassland and tillage land in WP1b, where nBTPT (250, 500 and 1000 ppm) reduced NH3 loss by a mean of 70% (range 25-100%).

Averaged over all rates of nBTPT, formulations and soil type, total NH3 loss was greatest at 15C and least at 5C (Table 2). Although total NH3 emission at 25C was lower than at 15C, the time of the maximum rate of NH3 loss (Tmax) was much earlier (Table 2). At 25C emission rates were high during the first few days but fell rapidly, probably because the soil dried out. Over all, total NH3 loss from the soils was in the following order DB>GL>HM>BX. Cation exchange capacity has been shown to be negatively correlated to NH3 loss from urea (Stevens et al., 1989), due to the fixation of ammonium-N ions on to soil cation exchange sites. In this study, there was not a good relationship between CEC and total NH3

volatilisation although, the DB and GL soils had lower CEC’s compared to HM and BX. This agrees with the results from the factors experiments in WP1b, which were unable to establish a relationship between NH3 loss and soil properties.


Table 1. Soil Properties

Soil pH Sand(%)

Silt(%)

Clay(%)

LOIa

(%)N

(%)Org C(%)

CECb

(meq kg –1)Bulk density

(g cm-3)MCc

(%)Olsen P(mg l –1)

Extd K(mg l –1)

Ext Mg(mg l –1)

Urease Activity(μg NH4-N g-1 hr-1)

BX 6.94 34.20 20.11 45.69 8.10 0.26 2.46 245 1.3 32.5 22 278 120 34.65

GL 6.45 82.80 12.15 5.05 2.70 0.08 1.01 48 1.3 14.1 57 140 66 6.16

DB 6.41 60.30 24.72 14.98 5.30 0.17 1.63 105 1.3 23.4 26 117 60 21.17

HM 6.36 21.59 50.95 27.46 9.20 0.34 3.31 215 1.1 33.5 12 126 97 48.93

a Loss-on-ignition, b Cation-exchange capacity, c moisture content at field capacity, d extractable

Table 2. Over all effect of temperature, formulation and soil type on soil moisture loss and NH3 emission.

Temperature (C) Formulation Soil5 15 25 S.E. Signif Coated Melted UAN S.E. Signif BX GL DB HM S.E. Signif

Moisture loss (%) 56.9a 67.7ab 98.3b 10.28 * 73.8 74.0 75.0 0.67 NS 69.4a 81.5c 78.8b 67.5a 0.77 ***

Total NH3-N

loss (%)2.8a 8.7b 4.1a 0.84 * 5.7a 6.3b 3.6c 0.18 *** 3.7a 5.3c 7.1d 4.7b 0.20 ***

Tmax$

(d) 11.7 7.6 3.2 0.27 *** 8.29 8.09 6.09 0.27 *** 6.19 8.09 8.22 7.46 0.31 ***

Max daily NH3-N

loss (%)0.35 1.31 1.27 0.04 *** 1.03 1.30 0.61 0.04 *** 0.62 0.91 1.50 0.89 0.05 ***

Inhibition (%) 83.2c 61.3a 74.1b 4.00 * 74.9b 78.5c 65.1a 0.86 *** 67.8a 68.8c 79.6a 75.2b 0.99 ***

***,* effects significant at P<0.001, <0.05 respectively NS= Not Significant $Tmax= Time of maximum rate of NH3 lossMain effects within a row followed by the same letter are not significantly differen

NT2605 Final Report WP3…Optimum use of Agrotain________________________________________________________________________________

Figure 1. Effect of nBTPT concentrations (ppm) on total NH3-N loss (as % of N applied) for four soil types (Boxworth, Gleadthorpe, DeBathe, High Mowthorpe) and three formulations of urea (Coated, Melted, UAN solution) at 5ºC (vertical bars are standard errors).


b) Melted 5 0C

05

101520253035

BX GL DB HMSoil

NH

3-N

lost

(as

% o

f N

appl

ied)

c) UAN 5 0C

05

101520253035

BX GL DB HMSoil

NH

3-N

lost

( as

% o

f N

appl

ied)

a) Coated 5 0C

05

101520253035

BX GL DB HMSoil

NH

3-N

lost

(as

% o

f N

appl

ied)

01002505007501000




a) Coated 15 0C

05

10152025

3035

BX GL DB HMSoil

NH

3-N

lost

(as

% o

f N

appl

ied)

01002505007501000

b) Melted 15 0C

05

101520253035

BX GL DB HMSoil

NH

3-N lo

st (

as %

of N

ap

plie

d)

c) UAN 15 0C

05

101520253035

BX GL DB HMSoil

NH

3-N

lost

(as

% o

f N

appl

ied)




a) Coated 25 0C

05

101520253035

BX GL DB HMSoil

NH

3-N

lost

(as

% o

f N

appl

ied)

01002505007501000

b) Melted 25 0C

05

101520253035

BX GL DB HMSoil

NH

3-N

lost

(as

% o

f N

appl

ied)

c) UAN 25 0C

05

101520253035

BX GL DB HMSoil

NH

3-N lo

st (

as %

of N

ap

plie

d)


Table 3. Over all effect of nBTPT on total NH3 loss, maximum daily rate of loss and time of maximum rate of loss (Tmax).

nBTPT (ppm)0 100 250 500 750 1000 S.E. Signifa

Total NH3

loss (% N applied)

13.5 5.2 3.9 3.3 2.6 2.7 0.25 ***

Max daily rate (%) 3.36 0.85 0.56 0.45 0.34 0.31 0.06 ***

T max (days) 4.2 7.0 7.7 7.9 8.8 9.4 0.38 ***

a ***P<0.001

Over all, total NH3 loss was significantly higher from the melted than from the coated product (Table 2). This effect was due primarily to a significant rate x formulation interaction, as shown in Fig 4. Ammonia loss from the 0 ppm nBTPT coated product was significantly lower than the melted product, which indicated that propylene glycol alone had a small effect in lowering NH3 loss from granular urea. Ammonia loss expressed as a % of the N applied was significantly lower from UAN compared to the granular products, as only 52.8% of the N applied in UAN was in the urea form.

If the NH3 loss was expressed as a % of the urea-N applied then the UAN treatment was significantly higher than the granular products. Averaged over all treatments, total NH3 loss expressed as a % of the urea-N applied was 6.8, 5.7 and 6.3 for the UAN, coated and melted products, respectively

Figures 5, 6 and 7 shows the percentage inhibition of total NH3 loss for each soil, formulation and rate of nBTPT at 5ºC, 15ºC and 25ºC, respectively. Averaged over all treatments, the %


Figure 4. Effect of nBTPT concentrations (ppm) on total NH3-N loss (% of N applied) (averaged over temperatures and soils)

LSD 1.189

0

4

8

12

16

20

Melted Coated UANFormulation

NH3-N

loss

(% o

f N

appl

ied)

01002505007501000


inhibition in NH3 loss was significantly lower at 15ºC (61%) than at 5ºC (83%) or 25ºC (74%) (Fig 8) and was significantly lower with UAN solutions (65%) compared to the granular formulations (75-79%) (Fig 9, Table 2). The results of field trials in WP1b also showed that the inhibitor was less effective in lowering NH3 emission from UAN solutions than with solid urea. Carmona et al. (1990) found that higher concentrations of nBTPT were required at 32ºC than at 18ºC to achieve equivalent suppression of NH3 loss. However, in this study there was no evidence that the % inhibition in NH3 loss at 15ºC or in UAN solutions was increased by higher concentrations of nBTPT. There was little benefit in increasing the nBTPT concentration above 250 ppm, with any of the formulations. Over all, the % inhibition with the melted products (79%) was significantly higher than with the coated material (75%), although the effect was small. Over all, the % inhibition was significantly higher with the 2 grassland soils (DB 80% and HM 75%) compared with the 2 arable soils (BX 68% and GL 69%) (Fig. 10, Table 2). This effect was predominantly due to a low % inhibition with the BX and GL soils at 25ºC with UAN (Fig 7). In a previous study with sixteen soils, Watson et al. (1994) showed that the response to increasing inhibitor concentration in lowering NH3

volatilisation was greatest in a soil with low organic matter content and high pH. The number of soils in the current study was too limited to confirm this observation.

It is not clear why the % inhibition with nBTPT was lower at 15ºC compared to the other two temperatures and why it was less effective in a UAN solution than in granular products. The effect may be due to the speed of formation of the oxygen analogue in soil and/or its stability. It is possible that at 15ºC and in UAN solutions, the rate of urea hydrolysis was more rapid than the rate of nBTPT conversion to its oxygen analogue. There is no information on the biotic and abiotic factors affecting the rate of conversion to NBPTO and its subsequent stability in soil. This is an important area for further investigation. Figure 11 shows that there was a significant soil x temperature interaction in the % inhibition by nBTPT, averaged over all rates and formulations, which supports the view that these factors may be important in the speed of formation of NBPTO.

4.2. Effect of nBTPT on soil N transformations

In the unamended urea treatment the % urea remaining at the end of the incubation averaged 16.3%. This was higher than expected, as evidence suggests that urea hydrolysis is rapid in moist soils (O’Toole and Morgan, 1988; Watson et al., 1994). The large amount of urea remaining in the unamended urea treatment probably reflected the high moisture loss of the soils, particularly at 25C. There was significantly more urea remaining and significantly less NH4

+-N in the nBTPT treatments as compared to the unamended treatment, which showed that nBTPT was delaying urea hydrolysis (Table 4).


Figure 5. Effect of nBTPT concentrations (ppm) on percent inhibition of NH3-N loss for each soil type (Boxworth, Gleadthorpe, DeBathe, High Mowthorpe) and formulation of urea (Coated, Melted, UAN solution) at 5ºC (Vertical bars are standard errors).

. a) Boxworth-5 0C

0

20

4060

80100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion

CoatedMeltedUAN

b) Gleadthorpe-5 0C

0

20

40

60

80

100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion

c) DeBathe-5 0C

020406080

100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion

d) High Mowthorpe-5 0C

020406080

100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion


.a) Boxworth-15 0C

0

20

40

60

80

100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion

CoatedMeltedUAN

b) Gleadthorpe-15 0C

0

20

40

60

80

100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion

c) DeBathe-15 0C

020406080

100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion


020406080

100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion


a) Boxworth-25 0C

020406080

100

0 100 250 500 750 1000ppm nBTPT

% in

hibi

tion

CoatedMeltedUAN

b) Gleadthorpe-25 0C

0

20

40

60

80

100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion

c) DeBathe-25 0C

0

20

40

60

80

100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion


0

20

40

60

80

100

0 100 250 500 750 1000ppm nBTPT

% In

hibi

tion


Figure 8. The effect of nBTPT concentration on percent inhibition in NH3-N loss at three temperatures (5, 15, 25ºC) (LSD= 13.36).

Figure 9. Effect of nBTPT concentration on percent inhibition in NH3-N loss in three formulations (Coated, melted and UAN solutions) (LSD= 5.34).

Figure 10. Effect of nBTPT concentration on percent inhibition in NH3-N loss from four soils, averaged over all temperatures and formulations (LSD= 6.17).


0

20

40

60

80

100

0 200 400 600 800 1000ppm nBTPT

% in

hibi

tion

51525

0

20

40

60

80

100

0 200 400 600 800 1000ppm nBTPT

% in

hibi

tion

Melted

Coated

UAN

0

20

40

60

80

100

0 200 400 600 800 1000

ppm nBTPT

% in

hibi

tion

BXGLDBHM


Table 4. Over all effect of nBTPT on soil N transformations.

% of applied N remaining as:

nBTPT (ppm)

0 100 250 500 750 1000 S.E. Signif

Urea 16.3 23.1 26.8 28.0 29.3 30.7 0.95 ***

NH4+-N 37.1 35.6 33.4 32.6 31.7 31.6 0.81 ***

NO3--N 21.4 21.8 21.1 20.8 21.2 20.0 0.65 NS

% recovery 88.2 85.6 85.2 84.6 84.8 85.0 0.81 ****,* effects significant at P<0.001, <0.05, respectively NS= Not Significant % recovery includes NH3 emission

Because urea can be taken up by plant roots as the intact molecule (Bollard et al., 1968; Harper, 1984), such persistence could have physiological implications for plants and is an area meriting further investigation. Over all, the urea remaining was higher in the GL and DB soils, where the % moisture loss was significantly greater, than with the BX and HM soils (Table 5).

The % of applied N remaining as urea was significantly lower with UAN than with the granular products (Table 5) as only 52.8% of the N applied was in the urea form to start with. If urea remaining was expressed as a % of the urea-N applied then there was no significant difference between UAN and the granular products.

The inhibitor had no significant effect on the % N applied remaining as NO3--N in the soil

(Table 4), agreeing with other workers that urease inhibitors had little effect on nitrification

NT2605 Final Report Optimum use of Agrotain.doc

Figure 11. Effect of temperature on percent inhibition in NH3-N loss from four soils, averaged over all nBTPT

concentrations and formulations (LSD= 13.21).

0

20

40

60

80

100

5 15 25Temperature 0C

% in

hibi

tion

BX

GL

DB

HM

23


(Bundy and Bremner, 1974; Bremner et al., 1986). Watson et al. (1994) showed that nBTPT at the 2,800 ppm level significantly lowered soil NO3

--N. However, this concentration was considerably higher than that used in the current study. There was a highly significant difference between soils in the % N applied remaining as NO3

--N (Table 5). With the BX soil, over all treatments, 36.7% of the N applied was as NO3

--N at the end of the incubation, which reflected high rates of nitrification. This soil was from an arable site and had the highest pH (6.9). A high soil pH has been reported to be one of the most important soil properties influencing nitrification (Morrill and Dawson, 1967; Haynes, 1986). Arable soils have been shown to have higher nitrification rates than grassland soils (Schmidt, 1982). Although the GL soil was also from an arable site, its limited nitrification rate was probably due to its high moisture loss. The % N applied remaining as NH4

+-N was significantly lower in the 2 arable soils (BX and GL) compared to the 2 grassland soils (DB and HM) (Table 5).

The low proportion of applied N remaining as NO3--N in soil at 25C (Table 5) probably

reflected the rapid moisture loss at this temperature. The proportion of applied N remaining as NO3

--N was also low at 5C, probably due to the sensitivity of nitrifying bacteria to low soil temperatures. This was supported by the fact that the % of the applied N remaining as NH 4

+-N was highest at 5 C.

Over all, there was no significant difference between the coated and melted granular fertilisers in the % of the applied N remaining as urea, NH4

+-N or NO3--N (Table 5). As

expected, NH4+-N and NO3

--N concentrations were significantly higher with UAN, as this treatment included both N forms. The total N recovered was calculated as total NH3

volatilised over 21 days plus the proportion of applied N recovered in the soil as NH4+-N,

NO3--N and urea. The unaccounted for N in the soils was presumably fixed, immobilised into

soil organic matter, lost by aerobic denitrification, or other gaseous emission.

Over all, the % N recovered was significantly higher with the unamended urea treatment (88.2%) compared with the treatments containing nBTPT (~ 85%, Table 4). This suggested that the inhibitor was having a small but significant (P<0.05) effect on either immobilisation or other soil N cycle processes. There was a small but significant difference between soils. Total N recovered was higher in the GL and DB soils, where the % moisture loss was significantly greater, than with the BX and HM soils (Table 5). Over all, the total N recovered was significantly higher (P<0.001) with UAN (91.2%), which was applied as a liquid, than either of the granular products (83.2% and 82.2% for the coated and melted products, respectively, Table 5). Further work is required to elucidate these effects. At 25 C the % N recovered was lower than at 5C or 15C. However, over all the effect of temperature on N recovery was not significant (P>0.05), due to high variability in the data.


Table 5. Over all effect of temperature, formulation and soil type on N transformations.

% of applied N remaining as:

Temperature C Formulation Soil

5 15 25 S.E. Signif Coated Melted UAN S.E. Signif BX GL DB HM S.E. Signif

Urea 23.9 17.0 36.2 10.08 NS 30.6a 29.4a 17.2b 0.67 *** 21.7c 33.9a 27.3b 19.9c 0.77 ***

NH4+-N 47.3a 31.4ab 22.2b 5.91 * 32.3b 32.1b 36.6a 0.57 *** 22.8d 30.7c 39.8a 41.4a 0.66 ***

NO3--N 17.7b 33.2a 12.3b 5.08 * 14.7b 14.5b 33.9a 0.46 *** 36.7a 16.5c 12.6d 18.5b 0.53 ***

Total N recovered (%) 91.7 90.3 74.6 5.0 NS 83.2b 82.2b 91.2a 0.57 *** 84.8b 86.3a 86.7a 84.4b 0.66 *

***, * effects significant at P<0.001, <0.05, respectively NS= Not Significant. Total N recovered includes NH3 emission Main effects within a row followed by the same letter are not significantly different


4.3. Stability of nBTPT during storage

Figure 12 shows the % recovery of nBTPT over a 17 month period in the three formulations stored in a fertiliser store (average temp. 11.4C). The nBTPT in the coated material degraded much more rapidly than in either the melted or UAN products.

The half-life of 1000 ppm nBTPT in the coated product was approximately 113 days, whereas the half-life in the melted and UAN products was similar, at 270 days. After 17 months storage the % recovery in the coated, melted and UAN products was similar at 25%, 28% and 27%, respectively.

Figure 13 shows the HPLC trace of the melted product on 2 occasions: 23 March and 6 August 2004. The nBTPT concentration was 1190 ppm on 23 March, which fell to 829 ppm on 6 August 2004, as indicated by the lower peak height at retention time 6.57 mins. No other major peaks were present in the vicinity of nBTPT. In contrast, with the coated product, as the nBTPT peak fell with time a second compound appeared with a retention time of 5.85 mins (Fig 14). The identity of this compound is unknown.


Figure 12. Percent recovery of 1000 ppm nBTPT in three formulations in the fertiliser store.

0

20

40

60

80

100

120

14/03/2004 07/06/2004 31/08/2004 24/11/2004 17/02/2005 13/05/2005 06/08/2005

Date

% re

cove

ry o

f nBT

PT

UANCoatedMelted


Figure 13. HPLC trace of 1000 ppm nBTPT in the melted product.

Figure 14. HPLC trace of 1000 ppm nBTPT in the coated product.

Clearly the decomposition of the melted and coated products was different. An important aspect in the degradation of the coated material is the identity of the unknown compound and whether it has an inhibitory effect on soil urease. Further work is required to elucidate these findings.

The nBTPT in the melted products maintained in bulk quantities at 4ºC was stable, at all concentrations (Fig 15). nBTPT in the coated products was much more stable at 4ºC (Fig 16) than in the fertiliser store at an average temperature of 10.3ºC, over the same time period (23 Mar 2004 – 27 May 2005).


6.57

6.57

Analysed 23/03/04

Analysed 06/08/04

6.57

Analysed 23/03/04

Analysed 06/08/04

5.856.57

Time

Time

1190 ppm nBTPT

829 ppm nBTPT

794 ppm nBTPT

249 ppm nBTPT


There was rapid deterioration in nBTPT at 4ºC when the fertiliser was placed in a 350 ml screw top plastic jar. The coated material was very deliquescent and after one months storage in the jar at 4ºC, the product was very slushy. Re-sampling the original bulk sample from a plastic bag, showed that the rate of degradation was much slower (Fig. 16). This study suggested that temperature and moisture might be important in the stability of nBTPT in coated urea products.


Figure 15. The stability of nBTPT in Melted products at 4 0C.

0

200

400

600

800

1000

1200

1400

14/01/04 23/04/04 01/08/04 09/11/04 17/02/05 28/05/05

Date

ppm

nB

TPT

1000

750500

250100

Figure 16. The stability of nBTPT in Coated products at 4 0C.

0100200300400500600700800900

14/01/04 23/04/04 01/08/04 09/11/04 17/02/05 28/05/05

Date

ppm

nB

TPT

1000

750500

250100

ppm nBTPT

ppm nBTPT

Stored in jars Stored in original bags


To investigate the temperature effect a further storage trial was established during 2005 at 4ºC, 15ºC and 25ºC. Samples were also stored in the fertiliser store for comparative purposes.

The ‘old melted’ material stored at 4ºC confirmed that nBTPT was reasonably stable at this temperature (Fig. 17). However, there was significant degradation of nBTPT at the higher temperatures, particularly at 25ºC. After 5 months, % recovery of nBTPT at 4ºC, 15C, fertiliser store (average temperature 13.8ºC) and 25ºC was 92%, 76%, 76% and 43%, respectively. The recovery of nBTPT in the ‘new melted’ product was considerably better than in the ‘old melted’ product, being 99%, 92%, 96% and 83%, after 5 months storage at 4ºC, 15ºC, fertiliser store (average temperature 13.8ºC) and 25ºC, respectively (Fig. 18). Although the new manufacturing procedure had improved the stability of nBTPT, there was still significant degradation at 25ºC.

The % recovery of nBTPT in the coated product was highly temperature dependent being 83%, 50%, 55% and 24% after 5 months storage at 4ºC, 15ºC, fertiliser store (average temperature 13.8ºC) and 25ºC, respectively (Fig. 19). The rate of degradation in the fertiliser store was not as rapid with the product coated on 11 January 2005 at 500 ppm nBTPT (half-life > 167 days) as with the coated product manufactured on 13 February 2004 at 1000 ppm nBTPT (half-life 113 days). The average daily temperatures from March to August 2004 and 2005 in the fertiliser store were very similar at 13.6 and 14ºC, respectively, so the effect does not appear to be due to differences in temperature. The main differences between the two coated products was the container used to store them in and the rate of nBTPT applied. In 2004, the product was stored in a screw top jar and had been coated with nBTPT at a theoretical rate of 1000 ppm. In 2005, the product was stored in a sealed plastic bag and coated at a rate of 500 ppm nBTPT. The plastic bag would have excluded moisture and air better than the screw top jar. However, when the 500 ppm coated product was vacuum packed, this did not improve the stability of nBTPT (Fig. 20), which suggested that moisture or air was not a major factor influencing degradation of nBTPT in this product. The rate of nBTPT application may explain the effect. The rate of degradation at 1000 ppm was higher than at 500 ppm. Either there is more self-catalysis occurring at the higher nBTPT concentration or the additional propylene glycol used to dilute the Agrotain solution (at 500 ppm rate) was having a beneficial effect. Clearly the stability of the coated products is very dependent on storage conditions, and further work would be required to fully elucidate these effects.

The % recovery of nBTPT in UAN solutions was also highly temperature dependent being 85%, 57%, 59% and 14% after 5 months storage at 4ºC, 15ºC, fertiliser store (average temperature 13.8ºC) and 25ºC, respectively (Fig. 21). The rate of degradation of nBTPT in UAN solutions stored in a fertiliser store was very similar in the 2004 and 2005 storage trials.



Figure 17: Percent recovery of ~500 ppm nBTPT in ‘Old Melted’ product stored at 4ºC, 15ºC, 25ºC and fertiliser store.

Figure 18: Percent recovery of 500 ppm nBTPT in ‘New Melted’ product stored at 4ºC, 15ºC, 25ºC and fertiliser store.


0

20

40

60

80

100

120

09/03/05 28/04/05 17/06/05 06/08/05 25/09/05

Date

% re

cove

ry o

f nB

TPT

4 C15 C25 CStore

0

20

40

60

80

100

120

09/03/05 28/04/05 17/06/05 06/08/05 25/09/05

Date

% re

cove

ry o

f nB

TPT

4 C15 C25 CStore


Figure 19. Percent recovery of 500 ppm nBTPT in the non-vacuum packed coated products stored at 4ºC, 15ºC, 25ºC and fertiliser store.

0

20

40

60

80

100

09/03/05 08/05/05 07/07/05 05/09/05

Date

% re

cove

ry o

f nB

TPT 4 C

15 C25 CStore

Figure 20. Percent recovery of 500 ppm nBTPT in the vacuum packed coated products stored at 4ºC, 15ºC, 25ºC and fertiliser store.

0

20

40

60

80

100

09/03/05 08/05/05 07/07/05 05/09/05

Date

% re

cove

ry o

f nB

TPT 4 C

15 C25 CStore

Figure 21. The recovery of 500 ppm nBTPT in UAN solutions at 4ºC, 15ºC, 25ºC and fertiliser store.

0

20

40

60

80

100

09/03/05 08/05/05 07/07/05 05/09/05

Date

% re

cove

ry o

f nB

TPT 4 C

15 C25 CStore



5. Conclusions

nBTPT was highly effective in lowering NH3 volatilisation from urea and delaying the time of maximum rate of loss. The average % inhibition over all soils, temperatures and formulations was 61.2%, 69.9%, 74.2%, 79.2% and 79.8% for the 100, 250, 500, 750 and 1000 ppm nBTPT concentration, respectively.

There was little additional benefit in using concentrations above 250 ppm nBTPT in any formulation.

Ammonia volatilisation from unamended urea varied with soil type and temperature and ranged from 8.2% to 31.9% of the N applied. It was not possible to explain the variation in NH3 emission by temperature or measured soil parameters.

Time of maximum rate of NH3 loss was temperature dependant, 25C> 15C> 5C. However, total NH3 loss was greatest at 15C, probably because the soil dried out faster at 25C.

There was little difference between the melted and coated granular products in reducing NH3 loss or in soil N transformations.

NH3 loss from the 0 ppm nBTPT coated product was significantly lower than the melted product, which indicated that propylene glycol (used for coating) alone had a small effect in lowering NH3 loss from granular urea.

The % inhibition with nBTPT was lower at 15C compared to 5C or 25C and was lower in a UAN solution than in granular products. It was suggested that this effect may be due to the speed of formation of the oxygen analogue in soil and/or its stability.

There was no evidence that higher concentrations of nBTPT were required at 15 C or in UAN solutions to achieve the equivalent % inhibition in NH3 loss at 5C and 25C and in granular products.

Significantly, more urea remained in the soil at the end of the incubation in the nBTPT treatments compared to the unamended treatment, which showed that nBTPT was delaying urea hydrolysis.

There was no evidence that the inhibitor had any effect on net nitrification rates in soil. However, it decreased total N recovery suggesting a small but significant effect on either immobilisation or other soil N cycle processes.

The stability of nBTPT in all fertiliser formulations was highly dependent on temperature.

The nBTPT in the melted products was stable when stored at 4C. However, there was a small but significant decrease in nBTPT concentration in the coated and UAN solutions stored at 4C.



nBTPT in all formulations degraded more rapidly when stored at 15C, 25C or at fluctuating temperatures in a fertiliser store, than at 4C.

nBTPT in the coated material degraded much more rapidly than in either the melted or UAN products when maintained at fluctuating temperatures in an unheated fertiliser store over 17 months (average temp. 11.4C).

Vacuum packing the coated material did not improve the stability of nBTPT, at any temperature, suggesting that moisture or air was not a major factor influencing degradation.

The manufacturing procedure used to produce the melted product influenced the subsequent stability of nBTPT.

In any further studies the melted product (nBTPT added to the urea melt) should be used in preference to the coated material and stored in bulk quantities under cool dry conditions.

UAN solutions containing nBTPT should be made up fresh or stored at a cool temperature (4C), where the degradation of nBTPT is predicted to be 15% after 5 months.



6. Future research requirements

Analytical techniques need to be developed to measure and quantify the products of nBTPT degradation in fertiliser formulations and soil.

Further work is required on the biotic and abiotic factors controlling the conversion mechanism of nBTPT to its oxygen analogue and its subsequent stability in soil.

Further work is required to elucidate the effect of temperature and soil moisture per se on the efficacy of nBTPT in lowering NH3 loss from urea.

Future work is required to investigate the effect of nBTPT on mineralisation/ immobilisation turnover in soil and on other soil N cycle processes.

Further work is required to investigate why the total N recovered was higher with UAN compared with the granular forms.

Further work is required to elucidate the factors influencing the stability of nBTPT in a coated product and whether the degradation products have urease inhibitory activity.

The stability of nBTPT amended urea when stored in bulk blends with P and K fertilisers requires investigation.

Further work is required to compare the stability of nBTPT in UAN solutions.

Because urea can be taken up by plant roots as the intact molecule, the persistence of urea treated with nBTPT could have physiological implications for plants and is an area meriting further investigation.

In deciding on the optimum incorporation rate of nBTPT a cost : benefit analysis is required.



7. References

Bollard, E.G., Cook, A.R. & Turner, N.A. (1968). Urea as sole source of nitrogen for plant growth. I. The development of urease activity in Spirodela oligorrhiza. Planta 83, 1-12.

Bran and Luebbe Application Notes (1989). TON and nitrite in water and seawater for Traacs 800 continuous flow analyser. Method No. 369-88E. Ammonia in water and wastewater for Traacs 800 continuous flow analyser. Method 418-91E. Bran and Luebbe, Brixworth, Northants, UK.

Bremner, J.M., McCarty, G.W., Yeomans, J.C. & Chai, H.S. (1986). Effects of phosphoroamides on nitrification, denitrification, and mineralization of organic nitrogen in soil. Communications in Soil Science and Plant Analysis 17, 369-384.

Bundy L.G. and Bremner J.M. (1974). Effect of urease inhibitors on nitrification in soils. Soil Biology & Biochemistry 6, 27-30.

Byrnes, B.H. & Freney, J.R. (1995). Recent developments on the use of urease inhibitors in the tropics. Fertilizer Research 42, 251-259.

Carmona, G., Christianson, C.B. & Byrnes, B.H. (1990). Temperature and low concentration effects of the urease inhibitor N-(n-butyl) thiophosphoric triamide (nBTPT) on ammonia volatilization from urea. Soil Biology & Biochemistry 22, 933-937.

Dampney P.M.R., Chadwick D., Smith K.A. and Bhogal A. (2004). The behaviour of some different fertiliser-N materials. Report for Defra project NT2603.

Harper, J.E. (1984). Uptake of organic nitrogen forms by roots and leaves. In: Hauck R.D. (ed) Nitrogen in Crop Production. pp. 165-170. ASA, CSSA, SSSA, Madison, WI, USA.

Haynes, R.J. (1986). Uptake and assimilation of mineral nitrogen by plants. In: Haynes R.J. (ed). Mineral nitrogen in the plant-soil system, pp. 303-378. Academic Press, Inc., London.

MAFF (Ministry of Agriculture, Fisheries and Food) (1986). The Analysis of Agricultural Materials. HMSO, London.

Martens, D.A. & Bremner, J.M. (1984). Effectiveness of phosphoroamides for retardation of urea hydrolysis in soils. Soil Science Society of America Journal 48, 302-305.

Morrill L.G. and Dawson J.E. (1967). Patterns observed for the oxidation of ammonium to nitrate by soil organisms. Soil Science Society of America Proceedings 31, 757-760.

Mulvaney, R.L. & Bremner, J.M. (1978). Use of p-benzoquinone and hydroquinone for retardation of urea hydrolysis in soils. Soil Biology & Biochemistry 10, 297-302.

Mulvaney, R.L. & Bremner, J.M. (1979). A modified diacetyl monoxime method for colorimetric determination of urea in soil extracts. Communications in Soil Science and Plant Analysis 10, 1163-1170.

O’Toole, P. and Morgan, M.A. (1988). Efficiency of fertilizer urea: the Irish experience. In Nitrogen Efficiency in Agricultural Soils (D.S. Jenkinson and K.A. Smith, Eds), pp. 191-206. Elsevier Applied Science, Barking, England.



Rowell D.L. (1994). In Soil Science: Methods and Applications. pp.92-95. Longman Scientific and Technical.

Schmidt E.L. (1982). Nitrification in soil In: Stevenson F.J. (ed) Nitrogen in agricultural soils. pp. 253-287. ASA, CSSA, SSSA, Madison, WI, USA.

Skalar SAN++ Analytical User Manual v 1.2 (2003). Ammonia in soil extract Catnr. 155-322w/r issue 011304/EK/99228315. Nitrate + Nitrite in soil extract Catnr. 461-322 issue 011304/EK/99228315. Skalar (UK) Ltd, Wheldrake, York, UK.

Stevens R.J., Laughlin R.J. and Kilpatrick D.J. (1989). Soil properties related to the dynamics of ammonia volatilisation from urea applied to the surface of acidic soils. Fertiliser Research 20, 1-9.

Tabatabai M.A. (1982). Soil enzymes. In Methods of Soil Analysis No. 9, Part 2. Chemical and Microbiological Properties, (A.L. Page., R.H. Miller and D.R. Keeney Eds) 2nd

edn, pp.903-947, Madison, WI: American Society of Agronomy, Soil Science Society of America.

Watson, C.J. (2000).Urease activity and inhibition – principles and practice. Proceeding No. 454. Publ., The International Fertiliser Society, York, UK. 40pp.

Watson, C.J., Miller, H., Poland, P., Kilpatrick, D.J., Allen, M.D.B., Garrett, M.K. & Christianson, C.B. (1994). Soil properties and the ability of the urease inhibitor N-(n-butyl) thiophosphoric triamide (nBTPT) to reduce ammonia volatilization from surface-applied urea. Soil Biology & Biochemistry 26, 1165-1171.



8. Appendicies

Appendix 1. The concentration of nBTPT in the Coated, Melted and UAN solution (averaged over all experimental runs).

* Not sufficient data to calculate standard deviation.

NT2605 Final Report Optimum use of Agrotain.doc

Mean SD

Coated

1000ppm 905 90.2

750ppm 678 45.8

500ppm 440 25.4

250ppm 224 19.8

100ppm 93 8.7

Melted

1000ppm 1205 17.0

750ppm 808 14.2

500ppm 401 9.2

250ppm 226 6.4

100ppm 74 3.5

UAN

1000ppm 1026 33.5

750ppm 766 31.4

500ppm 504 24.4

250ppm 239 14.4

100ppm 106* *

37


Appendix 2. The variability of nBTPT concentration in the granular products (melted and coated).

Target nBTPT(ppm)

QUB analyses ppm(23/02/04) SD

Independent lab. analyses ppm

(28/01/04)Melted

1000 1216 26 1204750 833 15 861

500 404 13 449

250 243 10 278

100 80 2 123

Coated

1000 834 96 933750 701 60 707

500 407 24 455

250 186 16 240

100 72 8 127



Appendix 3. The analytical variability of the granular products (melted and coated).

Target nBTPT(ppm)

QUB analyses ppm(23/02/04) SD

Melted

1000 ND$ -

750 ND -

500 409 10

250 243 7

100 81 2

Coated

1000 931 6

750 ND -

500 468 5

250 216 2

100 87 2

$ND = Not determined


Appendix 4. Daily temperature measurements in the fertiliser store from 24 March 2004 to 26 August 2005.

02468

10121416182022242628

Date

Dai

ly T

emp

(0 C)

Max TempMin Temp

Mean Temp

Mean is the average of hourly recordings

Appendix 5A. Effect of nBTPT concentration (0, 100, 250, 500, 750, 1000 ppm) on daily NH3 loss from each formulation of urea (Coated, Melted, UAN solution) at 5 0C on arable soils (Vertical bars are standard errors).

Boxworth - Coated 5 0C

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

appl

ied)

0

100

250

500

750

1000

Gleadthorpe - Coated 5 0C

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

appl

ied)

Boxworth - Melted 5 0C

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

app

lied)

Gleadthorpe - Melted 5 0C

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H3-

N lo

st (a

s %

of

N a

pplie

d)

Boxworth - UAN 5 0C

0.00.10.20.30.40.50.60.70.8

Days

Dai

ly N

H3-

N lo

st (a

s %

of

N a

pplie

d)

Gleadthorpe - UAN 5 0C

0.00.1

0.20.30.4

0.50.6

0.70.8

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H3-

N lo

st (a

s %

of

N a

pplie

d)

Appendix 5B. Effect of nBTPT concentration (0, 100, 250, 500, 750, 1000 ppm) on daily NH 3 loss from each formulation of urea (Coated, Melted, UAN solution) at 5 0C on grassland soils (Vertical bars are standard errors).

High Mowthorpe - Coated 5 0C

0.0

0.5

1.0

1.5

2.0

2.5

3.0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Daily

NH 3

-N lo

st (a

s %

of

N ap

plie

d)

DeBathe - Coated 5 0C

0.0

0.5

1.0

1.5

2.0

2.5

3.0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Daily

NH 3

-N lo

st (a

s %

of

N ap

plie

d)

0

100

250

500

750

1000

DeBathe - Melted 5 0C

0.0

0.5

1.0

1.5

2.0

2.5

3.0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Daily

NH 3

-N lo

st (a

s %

of

N ap

plie

d)

High Mowthorpe - Melted 5 0C

0.0

0.5

1.0

1.5

2.0

2.5

3.0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

appl

ied)

DeBathe - UAN 5 0C

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

app

lied)

High Mowthorpe - UAN 5 0C

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

DaysD

aily

NH 3

-N lo

st (a

s %

of

N a

pplie

d)

Appendix 5C. Effect of nBTPT concentration (0, 100, 250, 500, 750, 1000 ppm) on daily NH3 loss from each formulation of urea (Coated, Melted, UAN solution) at 15 0C on arable soils (Vertical bars are standard errors).


0

1

2

3

4

5

6

7

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

app

lied)

0

100

250500

750

1000


0

1

2

3

4

5

6

7

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

app

lied)


0

1

2

3

4

5

6

7

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

appl

ied)


0

1

2

3

4

5

6

7

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

appl

ied)

Boxworth - UAN 15 0C

0.00.20.40.60.81.01.21.41.61.82.0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H3-

N lo

st (a

s %

of

N a

pplie

d)


0.00.20.40.60.81.01.21.41.61.82.0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

DaysD

aily

NH

3-N

lost

(as

%

of N

app

lied)

Appendix 5D. Effect of nBTPT concentration (0, 100, 250, 500, 750, 1000 ppm) on daily NH 3 loss from each formulation of urea (Coated, Melted, UAN solution) at 15 0C on grassland soils (Vertical bars are standard errors).

DeBathe - Coated 15 0C

0123456789

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Daily

NH

3-N lo

st (a

s %

of

N ap

plie

d)

0

100

250

500

750

1000

DeBathe - Melted 15 0C

0123456789

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Daily

NH 3

-N lo

st (a

s %

of

N ap

plie

d)High Mowthorpe - Coated 15 0C

0

12

3

45

6

78

9

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

app

lied)


0123456789

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

app

lied)


0

1

2

3

4

5

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Daily

NH 3

-N lo

st (a

s %

of

N a

pplie

d)

DeBathe - UAN 15 0C

0

1

2

3

4

5

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

Days

Dai

ly N

H 3-N

lost

(as

%

of N

app

lied)

Appendix 5E. Effect of nBTPT concentration (0, 100, 250, 500, 750, 1000 ppm) on daily NH 3 loss from each formulation of urea (Coated, Melted, UAN solution) at 25 0C on arable soils (Vertical bars are standard errors).


0

1

2

3

4

5

6

7

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Daily

NH 3

-N lo

st (a

s %

of

N ap

plie

d)01002505007501000


0

1

2

3

4

5

6

7

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

appl

ied)

Boxworth - UAN 25 0C

0.00.20.40.60.81.01.21.41.61.82.0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Dai

ly N

H 3-N

lost

(as

%

of N

app

lied)


0

1

2

3

4

5

6

7

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

app

lied)


0

1

2

3

4

5

6

7

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

app

lied)


0.00.20.40.60.81.01.21.41.61.82.0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Dai

ly N

H 3-N

lost

(as

% o

f N a

pplie

d)

Appendix 5F. Effect of nBTPT concentration (0, 100, 250, 500, 750, 1000 ppm) on daily NH3 loss from each formulation of urea (Coated, Melted, UAN solution) at 25 0C on grassland soils (Vertical bars are standard errors).

Debathe - Coated 25 0C

0

24

68

1012

1416

18

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

app

lied)

0

100

250

500

750

1000

Debathe - Melted 25 0C

0

2

4

6

8

10

12

14

16

18

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Dai

ly N

H 3-N

lost

(as

%

of N

app

lied)

Debathe - UAN 25 0C

0.00.51.01.52.02.53.03.5

4.04.55.0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Dai

ly N

H 3-N

lost

(as

%

of N

app

lied)

High Mowthorpe - Coated 25 0C

02468

1012141618

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Daily

NH 3

-N lo

st (a

s %

of N

ap

plie

d)


02468

1012141618

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Dai

ly N

H 3-N

lost

(as

% o

f N

app

lied)


0.00.51.01.52.02.53.03.54.04.55.0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Days

Dai

ly N

H 3-N

lost

(as

%

of N

app

lied)

report for defra project nt2601 - defra, uk - science...

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