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FINAL REPORT

20100067

ASSESSMENT OF UREASE AND NITRIFICATION INHIBITORS FOR IMPROVING NITROGEN USE

EFFICIENCY AND YIELD IN FORAGE SEED PRODUCTION

Funded by: The Agriculture Development Fund

August 2015

Prepared by: University of Saskatchewan

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1. PROJECT TITLE AND ADF FILE NUMBER Assessment of Urease and Nitrification Inhibitors for Improving Nitrogen Use Efficiency and yield in Forage Seed Production: Project # 20100067

2. PRINCIPAL INVESTIGATOR

Fran Walley Department of Soil Science, University of Saskatchewan [email protected]

3. CO-INVESTIGATOR

Rich Farrell Department of Soil Science, University of Saskatchewan [email protected]

4. ABSTRACT

Enhanced efficiency fertilizers (EEFs) are intended to reduce nutrient losses to the environment, thereby increasing nutrient availability for plant production. One category of EEFs is nitrogen (N) fertilizer that is stabilized using compounds such as urease and nitrification inhibitors, both of which slow the release of N by controlling key microbially mediated processes. Our study, comprised of field, laboratory and growth chamber experiments, was conducted to assess the use of EEFs to improve N use efficiency (NUE) and yield in forage grass (i.e., hybrid bromegrass and timothy) seed production. Field experiments revealed the efficacy of EEF products varied depending on the forage crop and time of application. For example, fall application of EEFs were generally more beneficial, particularly in hybrid bromegrass stands—increasing dry matter and seed yields by 4 to11% and 15 to 22%, respectively, compared to urea alone. Conversely, spring applications either did not significantly affect final yields or resulted in decreased dry matter and seed yields. Use of EEFs apparently delayed N-transformations that could lead to N losses and potentially reduce the size of the plant-available N pool. Overall, use of the EEFs resulted in small increases in NUE (2 to 5%) compared to untreated urea. In general, however, differences between urea and the EEFs were not statistically significant (p > 0.05).

Laboratory and growth chamber studies revealed that urease activity associated with organic matter accumulation is an important factor controlling the potential for gaseous N losses via ammonia volatilization. Although high soil pH frequently is cited as a main factor controlling ammonia volatilization losses, laboratory incubation experiments revealed that high volatilization losses occurred on relatively acid soils when high levels of urease were present. Thus, in existing grass forage stands where grass residues are retained on the surface (i.e., are not burned in the early spring) high urease activity is likely to promote ammonia volatilization losses, irrespective of soil pH. Under these circumstances, application of an EEF containing a urease inhibitor may be warranted.

5. INTRODUCTION

Management of fertilizer nitrogen (N) for forage seed production remains a challenge with many unanswered questions for today’s producers. Of great concern are

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the N losses associated with fertilizer N application. Fertilizer N that is lost to the environment is not available for plant growth, and consequently nitrogen use efficiency (NUE) is reduced. Lower nutrient use efficiencies typically are reported for surface applied fertilizers (Mahli et al., 1998; 2004), resulting in lower yields and greater risks to the environment. Surface application of N fertilizers, and urea in particular, is particularly troublesome because subsequent N losses can occur via multiple routes including ammonia volatilization, and nitrous oxide (a particularly potent greenhouse gas) losses associated with both nitrification and denitrification.

Improving NUE has many direct agronomic and environmental benefits. For the producer, reducing N losses can mean improved N availability to support increased crop production. Alternatively, if NUE can be improved, N fertilizer inputs can be reduced accordingly, while still maintaining the production potential. From an environmental perspective, increasing NUE means reducing harmful N losses to the environment.

Current SMA guidelines for “Fertilizer Management for Seed Production of Perennial Forages” indicate that “fall application (of N) is recommended for many grass species”, largely because many grass species form reproductive tillers in the fall. This is supported by AAFC research that showed greatest yield increases for N applied directly after harvest (i.e., late summer to early fall). Indeed, many producers in the Parkland region prefer to fertilize in the fall because wet spring conditions often restrict access to forage stands until after significant vegetative growth has occurred—at which time N deficiencies already have limited the yield potential. Unfortunately, fall applications of N, particularly broadcast applications of urea (which is the common practice), are subject to both gaseous volatilization and denitrification losses. Moreover, these losses may be maximized when broadcast applications occur in the fall, particularly on land where the pH is high (typical of many Parkland soils). With many producers applying N rates of approximately 90-100 lb N/ac, these losses represent a significant economic loss to producers, and a significant (and potentially harmful) loss to the environment.

Current SMA guidelines recognize the possibility of gaseous N losses and indicate “the best N source for surface-broadcast application is ammonium nitrate (34-0-0), because it is highly soluble in water and readily moves with soil moisture to roots for rapid uptake” (http://www.agriculture.gov.sk.ca/Default.aspx?DN=85fce0f4-f256-4d9b-8a73-65096196eb10). Unfortunately, ammonium nitrate is no longer readily available and thus producers need a new strategy, and new information, to meet their NUE and consequent yield goals.

Our study was initiated to examine the use of urease and nitrification inhibitors, both alone and in combination, for reducing gaseous N losses, improving NUE, and achieving economically feasible yield goals in grass seed forage production. Specifically, the objectives of the study were:

1) To examine the potential agronomic and environmental benefits of using urease and nitrification inhibitors, and combinations of both, in forage biomass and forage seed production in the Parkland region of Saskatchewan.

2) To assess the economic feasibility of using urease and nitrification inhibitors for forage seed production in the Parkland region of Saskatchewan.

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The project included field experiments in which we examined both the agronomic and economic feasibility of using urease and nitrification inhibitors at four field sites in the Parkland region of Saskatchewan. Laboratory and growth chamber experiments further examined factors which influenced the degree to which various inhibitors were successful in reducing losses.

6. METHODS

Experiments were conducted as originally proposed. As discussed in previous reports, we did not use a Fourier Transform Infra-Red trace gas analyzer (FTIR-TGA) connected to vented chambers to analyze ammonia (NH3) and nitrous oxide (N2O) emissions due to unsatisfactory results associated with the NH3 emissions. The gas sampling protocols that we chose for both field and growth chamber experiments were a suitable and scientifically accepted substitute.

Field Experiments

Four experimental sites were established within existing commercial forage seed production stands in the Boreal Transition zone of Saskatchewan near Carrot River (sites CR1 and CR2), Arborfield (ABR) and Choiceland (CHL). Both Carrot River sites were classified as Gleyed Dark Gray Chernozems and located within the same field in which a natural divide was identified such that CR1 (53o 12’41”N, 103 o 30’21”W) had higher pH values (pH 8.3) and lower organic matter levels than CR2 (53 o 12’26”N, 103 o 30’41”W) (Tables 1 and 2). The Carrot River sites were seeded to hybrid bromegrass (Bromus riparius Rehm. x Bromus inermis Leyss. cv. Success) in 2010. Timothy (Phleum pretense L. cv. Comer) stands were seeded at ABR in 2009 and CHL in 2011. The ABR site (53 o 10’52”N, 103 o 27’53”W) was classified as a Gleyed Dark Grey Chernozem and CHL (53 o 28’49”N, 104 o 29’22”W) was classified as a Dark Grey Chernozem.

Table 1. General characteristics of experimental sites used for the field experiments in 2012 and

2013.

Site Crop Stand establishment

Soil classification Bulk Density (g cm-1)

Organic Matter

(%) Carrot River 1 (CR1)

Hybrid bromegrass cv. Success

2010 Gleyed Dark Grey Chernozem

1.29 3.92

Carrot River 2 (CR2)

Hybrid bromegrass cv. Success


1.15

6.47

Arborfield (ABR)

Timothy cv. Comer


1.23 6.10

Choiceland (CHL)

Timothy cv. Comer

2011 Dark Grey Chernozem

1.13 6.15

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The experiments were conducted using a randomized complete block design with nine treatments replicated four times. Treatment plots measured 10.5 m wide x 11.2 m long with a 1 m margin between all plots and replicates. Prior to fertilizer application soil samples were collected from six random locations within each site at 0 to 15-, 15 to 30- and 30 to 45-cm depths. Samples were bulked, dried and sent to ALS Laboratories (Saskatoon SK, Canada) where extractable nutrients (N, P, K, S and Cu), total organic carbon, organic matter, pH and EC (1:2 soil water extraction) were determined (Table 2).

Treatments included an unfertilized control (C) with the remaining eight treatments representing the fall or spring application of four different enhanced efficency urea fertilizer (EEFs) products applied at a rate of 92 kg N ha-1. The four fertilizer treatments were 1) urea alone (U); 2) urea coated with the nitrogen stabilizer N‐(n‐butyl)thiophosphorictriamide(NBPT)(tradenameAGROTAIN® Koch Agronomic Services LLC, Wichita, KS, USA) (Ag) at the recommended rate of 1.5 g kg-1 urea; 3) urea containing both NBPT + dicyandiamide(DCD) (trade name SuperU™, Koch Agronomic Services LLC, Wichita KS, USA) (Su); and 4) urea containing both DCD + Triazole (trade name ALZON®, SKW Piesteritz, Wittenberg, Germany) (Al) (Table 3). Products were applied during the fall (September, 26nd and 27th, or all sites) of 2012 and the spring of 2013 (May 15th at ABR and CHL, and May 22nd at CR1 and CR2). A gaseous emissions sampling area was established in the center of each plot; these areas were covered with small pieces of plywood that were slightly larger than the footprint for the ammonia volatilization and nitrous oxide chambers to prevent broadcast fertilizer application on these areas. Fertilizer was surface applied to the remaining plot using a 1.7 m wide ground-driven fertilizer spreader pulled by an ATV to mimic a typical application method in these production systems.

Table 2. General soil chemical characteristics at the initiation of the field experiments

(September, 2013)

Site Depth (cm)

N‡ P# K# S‡ Cu¶ pH†† EC§ (dS m-1)

mg kg-1

Carrot River 1 0–15 8.9 17.9 130 15.6 0.62 8.3 0.226 15–30 3.8 6.2 60 22.0 0.49 8.3 0.234 30–45 3.6 < 2.0 51 44.5 0.58 8.4 0.265 Carrot River 2 0–15 9.0 6.7 106 8.9 0.69 7.5 0.203 15–30 5.9 2.7 64 6.8 0.64 8.1 0.174 30–45 4.6 < 2.0 58 7.1 0.16 8.2 0.158 Arborfield 0–15 9.7 11.6 302 6.3 0.90 6.2 0.050 15–30 6.7 4.5 186 6.9 1.1 6.9 0.074 30–45 7.4 2.2 151 12.3 0.23 7.7 0.231 Choiceland 0–15 6.1 6.0 152 25.9 0.50 7.4 0.237 15–30 3.1 < 2.0 67 44.3 0.43 8.2 0.234 30-45 1.7 < 2.0 66 62.3 0.38 8.0 0.276 ‡ Calcium chloride extractable. # Modified Kelowna extractable. ¶ DTPA extractable. ††pH of a 1:2 (soil:water) extract.

§ EC (electrical conductivity of a 1:2 (soil:water) extract.

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Table 3. Treatment used in the 2013/14 field experiments at Carrot River (1 & 2), Arborfield and

Choiceland. All fertilizer treatments applied to deliver 92 kg N ha-1.

Treatment Application Mechanism (inhibitor) Formulation

Control (unfertilized)

--

--

--

Untreated urea Fall 2012 None Spring 2013

AGROTAIN® Fall 2012 Spring 2013

Urease N-(n-butyl) thiophosphoric triamide (NBPT) (surface applied)

SuperU™ Fall 2012

Spring 2013 Urease & nitrification NBPT + dicyandiamide (DCD)

(incorporated) ALZON® Fall 2012 Nitrification DCD + Triazole (incorporated) Spring 2013 Gaseous nitrogen emissions

Nitrous oxide emissions were determined at all four sites, whereas ammonia volatilization emissions were determined at CR1 and CR2 only. Dates of fertilizer application and emission measurements are reported in Table 4. Base frames for emission chambers were inserted into the soil within the center of each treatment plot and remained in place during the measurement season. Fertilizer granules were applied by hand to the soil surface within the base frames either in the fall of 2012 or in the spring of 2013 to ensure that each chamber received N at a rate of 92 kg N ha-1. Ammonia and N2O emissions from fall-applied fertilizers were monitored immediately after fertilizer application until snowfall in 2012, and continued immediately after snowmelt in 2013. Emissions from spring-applied fertilizers were monitored immediately after application and continued until gas fluxes were negligible. Due to a high precipitation event following fertilizer application in the fall of 2012 and the low measured emissions, fertilizer application and monitoring of gaseous N emissions were repeated in the fall of 2013 at CR1 and CR2.

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Table 4. Time of fertilizer application and gaseous N emission measurements for the 2012-13 field experiments.

Site Fertilizer

application NH3 emission measurement

duration N2O emission measurement

duration CR1 10/8/2012 9/8/2012 – 18/8/2012 4/8/2012 – 19/8/2012 CR2 10/8/2012 9/8/2012 – 18/8/2012 4/8/2012 – 19/8/2012 ABR 10/8/2012 - 4/8/2012 – 19/8/2012 CHL 10/8/2012 - 4/8/2012 – 19/8/2012 CR1 10/8/2012 7/05/2013 – 14/05/2013 6/05/2013 – 27/05/2013 CR2 10/8/2012 7/05/2013 – 14/05/2013 6/05/2013 – 31/05/2013 ABR 10/8/2012 - 6/05/2013 – 27/05/2013 CHL 10/8/2012 - 7/05/2013 – 27/05/2013 CR1 23/05/2013 15/05/2013 – 3/06/2013 23/05/2013 – 12/06/2013 CR2 23/05/2013 15/05/2013 – 3/06/2013 23/05/2013 – 12/06/2013 ABR 15/05/2013 - 16/05/2013 – 20/06/2013 CHL 15/05/2013 - 16/05/2013 – 20/06/2013 CR1 18/09/2013 18/09/2013 - 25/09/2013 18/09/2013 - 8/10/2013 CR2 18/09/2013 18/09/2013 - 25/09/2013 18/09/2013 - 8/10/2013

Nitrous oxide emissions were estimated using vented emission chambers according to Yates et al. (2006). The system consisted of a circular PVC base frame and a vented cap fitted with a rubber sampling port. The chamber had a 0.02 m2 footprint and a closed chamber headspace of 2.25 L. The base frames were inserted into the soil prior to treatment application and remained in place throughout the sampling season, whereas the lid was placed on the base only during sampling events (Fig. 1).

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Figure 1. Installed base frame with sampling chamber lid, used to estimate nitrous oxide

emissions.

Gas samples were collected from the closed chamber at 10, 20 and 30 min using a 25-mL syringe with a 25-gauge needle, and were injected into 12 mL ExetainerTM tubes (Labco Limited, UK). An additional six to eight ambient air samples were drawn in pairs before treatment samples were collected at each site and the average was regarded as t0. Samples were transported back to the lab and the N2O concentration determined using a Bruker 450 GC gas chromatograph (Bruker Biosciences Corporation USA) according to Farrell and Elliott (2007). Fluxes were calculated by fitting either an exponential or linear regression through the concentration versus time data. Paired ambient air samples were used to calculate the minimal detectable concentration difference (MDCD) according to Yates et al. (2006). When concentrations of subsequent measurement intervals did not exceed the MDCD, they were regarded as not significantly different from each other and a linear regression was fitted to the data. When the concentration of subsequent samples exceeded the MDCD, a modified Hutchinson Mosier model (Hutchinson and Mosier, 1981; Pedersen et al., 2010; Pedersen, 2015) was fitted through the data. Fluxes were taken as the slope of either the linear regression or the Hutchinson Mosier model at t0. Cumulative emissions were calculated by interpolating flux values of adjacent sampling points (Pennock et al., 2006).

Soil NH3 emissions were estimated using a closed dynamic chamber system

consisting of a rectangular acrylic base frame (46 cm length x 22 cm width x 15.25 cm height) of 6.35 mm thickness and a lid containing a battery-powered air pump and a fan (Fig. 2).

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Figure 2. Closed dynamic chamber system used to estimate ammonia volatilization

emissions. The base frame (a) was installed to a depth of 5.25 cm (b) and left in the field for the duration of the sampling period. During emission measurements, the insulated lid (c) fitted with batteries (d) and an internal fan (e) attached to the baffles (f) and a sampling port (g) was attached to the base frame. Air from the headspace passed through tubing (h) leading to a sulfuric acid trap (i), which then returned the air to the chamber (j). Adhesive-backed foam stripping (k) preventing air exchange between the lid and the base.

The chamber bases were installed into the soil to a depth of 5.25 cm, resulting in a

headspace of 10.12 L. When the lid was fastened to the base, the inlet and outlet of the lid were connected to a vial containing 35 mL of a 0.5% H2SO4 solution. When the air pump and fan were activated, the air in the headspace of the chamber was guided through the H2SO4 solution at a speed of 2 L min-1, thereby trapping NH3 in the acid solution while NH3-free air was returned into the chamber. The lid was only fastened to the base during sampling and was removed between sampling times. In the fall of 2012, the sampling duration was limited to 30 minutes per day, largely because the batteries operating the fan system had a limited lifetime, particularly under cold conditions. After the fall measurements were completed, the system was modified by changing to a more energy-efficient pump and increasing the number of batteries powering the system. Emissions in all subsequent measurement seasons (i.e., spring 2013 and fall 2013) were measured for 90 min.

Measurements were started daily at 12:00 pm on one site and at 2:00 pm on the other, and the starting site was switched daily to minimize the impact of measurement timing on emission patterns. After the measurement, the H2SO4 solutions were transferred into 50 mL Falcon tubes and transported back to the lab for subsequent analysis. Concentration of NH3 within the acid solution was determined colorimetrically

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using a SmartChem 200 autoanalyzer (Westco Scientific Instruments Inc., CT, USA). Fluxes were calculated as the mass of NH3-N captured in the acid trap divided by the duration of sampling. Cumulative emissions were calculated as the area under the curve between fluxes of all sampling points, which assumes that emission rates remained constant throughout the sampling day.

Soil moisture determination

Volumetric soil moisture content (VMC%) was measured on each sampling day from within experimental plots at sites CR1 and CR2 using a portable TDR field probe. Within each plot, seven consecutive measurements were taken at a distance of approximately 15 cm from the emission chambers, and the values were averaged.

Biomass sampling

Biomass samples were collected at 14, 28, 42 and 71 d after fertilizer application at the timothy sites and at 14, 28 and 42 d at the hybrid bromegrass sites. Samples were taken using a 1-m2 wire frame placed within the plot at a random location. At seed maturity, which occurred 91 d after spring fertilizer application for timothy and at 70 d for hybrid bromegrass, sampling size was increased to 2 m2. Frames were placed away from the plot edges, with adequate separation from past biomass samples to eliminate edge effects. Sampled areas were flagged to avoid resampling at later dates. Biomass was cut by hand as close to the soil surface as possible using a sickle. The samples were dried at 60oC, weighed and subsampled. Subsamples were ground to pass a 2 mm screen using a Thomas Scientific Model 4 Wiley mill (Thomas Scientific, Swedesboro NJ, USA). All samples were analyzed for N using a LECO TruMac CNS (LECO Corporation, St. Joseph MI, USA).

Seed Yield

Final biomass samples were threshed after drying using a stationary threshing machine. Seed was cleaned using a series of hand sieves ranging from 2.0 to 0.5 mm to remove as much residue as possible while preserving total seed yield. After cleaning, samples were weighed to determine yield. Seed counts were conducted by hand and 1000 seed weight were determined based on 100 seed counts. Seeds were subject to H2SO4/H2O2 digestion (Thomas et al., 1967) and N was determined colorimetrically using a SmartChem@200 discrete analyzer (Westco Scienitific Instruments, Brookfield CT, USA).

Economic Analysis

The economic analysis is based upon fertilizer and product prices as of the fall 2013. No price premium was built into retail prices for spring or fall delivery to avoid skewing economic returns due to external factors. As ALZON® currently is not available for sale in Canada, prices were obtained from Gleadell Agriculture (Lincolnshire, UK).

Statistical Analysis

Statistical analysis was performed using R version 3.03 (R Foundation for Statistical Computing, 2014) using the nlme (Pinheiro et al., 2014) package with Tukey HSD post-hoc tests for agronomic data.

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Controlled Environment Experiments

Ammonia Volatilization Chamber Experiments

A series of experiments were conducted using ammonia volatilization chambers to assess the impact of pH, soil moisture and temperature on ammonia volatilization from various fertilizer sources (i.e., untreated urea versus various EEFs). Ammonia volatilization cabinets (Fig. 3) were constructed according to Woodward et al. (2011). The cabinets consist of a closed airflow system through which air is pumped through two sets of humidifiers (one outside, and one within the temperature-controlled cabinets) before it enters the soil chambers (containing 300 g of air-dry equivalent soil). Air exiting the soil chambers is then forced through acid traps containing 200 mL of 0.01 M H2SO4 thereby trapping the ammonia. Trapped ammonia was subsequently measured using a SmartChem 200 autoanalyzer (Westco Scientific Instruments Inc., CT, USA). The volatilization cabinets are insulated and temperature-controlled through a heating cable and a fan.

Air-dried and sieved soils used in the volatilization chamber experiments were brought to the desired moisture content prior to the initiation of each experiment and allowed to incubate for 7 d at 26o C to re-establish microbial communities. Immediately after incubation, the soils were sampled and initial soil ammonium and nitrate was determined. Soils were transferred to the soil chambers and treatments were surface applied. To examine the impact of soil pH on ammonia volatilization, soils were treated with CaCO3. Typically, experiments were conducted over 14 d, after which soils were subsampled and final ammonium and nitrate levels were determined.

In addition to the EEF products used in the field experiments, an additional product, Piazur® was included. This fertilizer product contains the urease inhibitor 2-nitrophenyl phosphoric triamide (2-NPT) and differs slightly from NBPT used in AGROTAIN®, but has the advantage that the inhibitor is incorporated into the fertilizer granules in a manner similar to SuperU™ and Alzon®. Enhanced efficiency fertilizers were tested in eight volatilization experiments with the soil pH ranging from 5.86 to 7.75, soil moisture content ranging from 50% of field capacity to 100% of field capacity, and soil temperature ranging from 5°C to 26°C.

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Figure 3. Ammonia volatilization chambers used in laboratory incubation experiments. Assessment of gaseous N losses using 15N labeled urea

A final controlled environment using 15N labeled urea together with the inhibitors was conducted to examine the fate of applied urea (i.e., gaseous N emissions and plant N uptake). Gaseous emissions were measured using the same ammonia volatilization chambers that had been used in field experiments to assess ammonia volatilization (Fig. 2). The chamber lids were modified to accommodate a sampling port for N2O emissions. Wheat (Triticum aestivum L.) was used as a test crop as forage grasses are slow to establish, and seedling forage grasses are unlikely to represent N uptake in established grass stands. The experimental units consisted of plastic soil trays with dimensions nearly matching those of the field chamber ammonia emission bases (46 x 22 x 15 cm), which held a soil:sand mix (50:50 by weight) (Fig. 4).

Soil used in this experiment was collected from CR1 and mixed with acid washed silica sand. Field capacity of the mix was determined. The soil mix was brought to 75% field capacity and incubated for one week prior to seeding. The soil mix (7.75 kg oven-dry equivalent) was placed in the trays to a depth of approximately 10 cm and the ammonia volatilization field chambers were inserted into the soil mix to a depth of 5 cm. Wheat was seeded into the incubated soils 7 d prior to the initiation of the experiment (i.e., fertilizer addition) in two rows at 15 cm row spacing.

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Figure 4. Experimental units used to assess gaseous losses from 15N-labelled urea under controlled growth chamber conditions.

Four N fertilizer treatments were as follows: 1) untreated 15N urea (10% 15N excess) added as a solution at a rate equivalent to the field rate (i.e., 92 kg N ha-1), or 15N labeled urea together with 2) AGROTAIN; 3) DCD (Sigma-Aldrich Co.); 4) a mixture of both AGROTAIN and DCD. Inhibitor amendments were added to the urea solution at rates equivalent to those used in the field. The fertilizer solutions were applied to the surface of the soil in droplets using a pipette, and droplets were evenly distributed to simulate fertilizer granule placement. The moisture content of the soil mix was maintained at approximately 75% field capacity by additions of water to the soil surface, as required. Gaseous emissions were assessed 24 h after fertilizer application and every 24 h thereafter for 10 d. At each sampling time, both N2O and NH3 emissions were determined. Nitrous oxide emissions were collected through a sampling port installed in the lid of the chamber. Ambient air samples were taken at T0 (i.e., immediately after the lids were closed) at each sampling time.

At 24 h intervals, lids were attached to the chamber bases and after 30 min, two gas samples were collected. The first was analyzed on a gas chromatograph to determine total N2O, and the second sample was analyzed to determine 15N2O using a Picarro G5101-i isotopic N2O analyzer. Ammonia emissions were determined after 30 min incubation, as

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described for the field experiments. In addition to the collecting ammonia in the acid traps, filter disks soaked in KHSO4 were installed within the chamber for the duration of the sampling period to trap 15NH3 labeled ammonia. The disks were encapsulated and analyzed using an isotope ratio mass spectrometer (Delta VAdvantage, Thermo Fisher Scientific Inc., Waltham, MA) coupled to an elemental analyzer (Costech ECS4010. Costech Analytical Technologies, Inc., Valencia, CA).

7. RESEARCH ACCOMPLISHMENTS

Objective Progress 1) To examine the potential agronomic and environmental benefits of using urease and nitrification inhibitors, and combinations of both, in forage biomass and forage seed production in the Parkland region of Saskatchewan.

Accomplished through fall and spring application of enhanced efficiency fertilizers, and measurement of forage grass biomass and final forage grass seed yields, and nitrous oxide and ammonia volatilization fluxes following fertilizer application at four commercial field sites in the Parkland region.

2) To assess the economic feasibility of using urease and nitrification inhibitors for forage seed production in the Parkland region of Saskatchewan.

Accomplished by assessing the returns associated with the various fertilizer treatments relative to the cost of applying the EEF versus urea alone.

8. RESULTS AND DISCUSSION

Field Experiments

Nitrous Oxide Emissions from Fall-applied Nitrogen Fertilizer

Nitrous oxide emissions were estimated prior to fall fertilizer application to establish baseline emissions, with regular monitoring initiated immediately after fertilizer application until snowfall in 2012 at all four sites. Emissions in the fall following fertilizer application were very low at all four sites (Figs. 5 to 7) and although the control (i.e., unfertilized) typically had the lowest emission rates, treatment differences were not significant. Excessively wet conditions following the fall 2012 fertilizer applications, necessitated that the fall applications be repeated in 2013, to confirm these observations.. Nitrous oxide emissions remained low and within a similar range (i.e., less than 25 g N ha-1 d-1) observed in the fall of 2012 (Figs. 8 and 9). A slight increase in emissions occurred following a precipitation event that enhanced soil water content. Although the emissions rates were low, data suggest that SuperU™, in particular, was associated with N2O emissions similar to the unfertilized control, whereas the AGROTAIN® treatment had emissions similar to the untreated urea. The relatively low emission rates in the fall of both 2012 and 2013 indicate that fall emissions generally did not pose a significant N loss potential, particularly if fertilizer application occurred late in the fall close to freeze-

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up. Importantly, the low rates of emissions, and the relatively short time frame over which emissions occurred suggest that although N2O losses may be of environmental significance, the losses were of little agronomic significance.

Nitrous oxide emissions were again monitored in the spring of 2013 for all fall-applied fertilizer treatments (Figs. 5 to 7) as soon as it was possible to access the field sites. At all four sites, spring emissions rates were lowest from the unfertilized control. Additionally, data suggest that early spring nitrous oxide emissions were influenced by temperature, with peak spring emissions occurring during periods of higher temperature.

Emissions at the first sampling date were elevated relative to fall emissions at CR1, CR2 and ABR, whereas the emissions at CHL for the first two sampling dates were similar to those obtained in the fall (Figs. 5 to 7). It is important to note that CHL was sampled 4 d after all other sites due to inaccessibility to the site, and it is likely that the early burst of emissions captured at the other sites was missed at CHL. Other researchers have reported high rates of N2O emissions during snowmelt (Yates et al., 2007).

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Figure 5. Nitrous oxide and ammonia fluxes in the fall of 2012 and spring of 2013 from

fall-applied N fertilizers at Carrot River 1 (CR1). Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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Figure 6. Nitrous oxide and ammonia fluxes in the fall of 2012 and spring of 2013 from

fall-applied N fertilizers at Carrot River 2 (CR2). Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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Figure 7. Nitrous oxide fluxes in the fall of 2012 and spring of 2013 from fall-applied N

fertilizers at Arborfield (ABR) and Choiceland (CHL). Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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Figure 8. Nitrous oxide and ammonia fluxes from fall-applied N fertilizers at Carrot

River 1 (CR1) in 2013. Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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Figure 9. Nitrous oxide and ammonia fluxes from fall-applied N fertilizers at Carrot

River 2 (CR2) in 2013. Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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The absolute ranking of the EEF treatments at the first spring sampling time is difficult to interpret because data suggest that the first sampling time may not have captured the peak early emissions. At both CR1 and CR2, the highest spring emissions were associated with the untreated urea, with emissions rates as high as 317 g N ha-1 d-1 detected at CR2 (Fig. 6). Thus, although AGROTAIN® and SuperU™ had higher emissions at ABR than untreated urea (Fig. 7), it is possible that emissions from the urea treatment occurred at an earlier date, and our measured estimates during the early sampling period reflect the end of peak emissions.

Spring N2O fluxes from fall applied N fertilizer additions, including EEFs, are an important finding. These spring losses are evidence that N losses from fall-applied urea-based fertilizers, and EEFs, can occur even if significant rainfall events follow fertilizer application in the fall. Indeed all sites experienced several minor rain events and one significant (>2.5 cm) event following fall fertilizer application in 2012. One might expect these rain events would be sufficient to move the fertilizer N into the soil, thereby protecting it from subsequent gaseous loss. Nonetheless, despite the fall precipitation, spring losses of from fall applied treatments were still noted suggesting that irrespective of any protection offered by the inhibitor treatments during the fall, the fall-applied N fertilizer remained subject to losses during early spring melt.

Ammonia emissions from fall-applied nitrogen fertilizer

Ammonia emissions were monitored at both CR1 and CR2 (Fig. 5 to 6). Ammonia emissions at CR1 were relatively low and unaffected by any of the treatments relative to the unfertilized control. At CR2, untreated urea was associated with the highest ammonia emissions, although relatively high variability may have obscured treatments effects.

Nitrous oxide emissions from spring applied nitrogen fertilizer

Nitrous oxide emissions from the untreated control (i.e., no fertilizer addition) remained very low in the spring of 2013 at all four sites, suggesting that available soil N levels contributed little to N2O losses (Figs. 10 to 12). Spring application of N fertilizer resulted in relatively rapid gaseous losses of N as N2O from the various treatments, with losses differing between treatments and sites. For example, at CR1, N2O losses were greatest from the untreated urea treatment with a spike in emission rates occurring approximately 5 d after application (Fig. 10). The ALZON® and AGROTAIN®

treatments followed a similar pattern, although emission rates were less than those observed for the untreated urea. SuperU™ provided the best control at CR1, with N2O emissions similar to the unfertilized control. In contrast, emission rates were similar between all treatments at CR2 (Fig. 11), and N2O emissions apparently were promoted in AGROTAIN® and SuperU™ treatments at both ABR and CHL (Fig. 12). This apparent enhancement of N2O losses was unexpected; however, although N2O losses were enhanced, losses remained very low at all sites, and typically did not exceed 100 g N ha-1 d-1, even during peak emissions. Thus treatment differences in N2O emissions are very small and likely not of agronomic significance. The relatively low emission rates likely reflect the low soil moisture content and the limited precipitation during the early spring of 2013. Others have reported that N2O emissions are highly sensitive to soil moisture content, with maximum emissions occurring when volumetric soil moisture in the range

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of 70 to 80%–values that typically are achieved following a rainfall event (Yates et al. 2007).

Figure 10. Nitrous oxide and ammonia fluxes in the spring of 2013 from spring-applied

N fertilizers at Carrot River 1 (CR1). Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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Figure 11. Nitrous oxide and ammonia fluxes in the spring of 2013 from spring-applied

N fertilizers at Carrot River 2 (CR2). Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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Figure 12. Nitrous oxide fluxes in the spring of 2013 from spring-applied fertilizers at

Arborfield (ABR) and Choiceland (CHL). Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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Ammonia emissions from spring applied nitrogen fertilizer

Ammonia emissions from spring-applied fertilizer N were significantly higher in the spring of 2013 compared to ammonia emissions from fertilizers applied in the fall of 2012. Specifically, maximum emissions at CR1 approached 590 g N ha-1 d-1 from untreated urea (Fig. 10) whereas ammonia emissions typically did not exceed 100 g N ha-

1 d-1 when measured following fall fertilizer application. Importantly, maximum volatilization losses occurred at CR1, where soil pH values were considerably higher (pH 8.3) than at CR2 (pH 7.5) (Fig. 11), where emissions more closely resembled those measured following fall application. The apparent impact of soil pH is consistent with many studies that have reported enhanced volatilization at high soil pH (Sommer et al., 2004).

Biomass Accumulation

Biomass accumulation was significantly enhanced by the application of fertilizer N relative to the unfertilized control at all sites (Fig. 13). Hybrid bromegrass was particularly responsive to N at both CR1 and CR2 whereas the magnitude of response of timothy to N at ABR, in particular, was less notable. Timothy biomass responses were greater at CHL than at ABR, likely reflecting the differences in the age of the stand. Arborfield, the relatively nonresponsive site, was seeded in 2009 whereas CHL was seeded in 2011. Although the magnitude of response to N fertilizer was less in timothy than in hybrid bromegrass, timothy was apparently highly dependent on fertilizer N for biomass production, with harvested biomass remaining largely unchanged over the growing season in the non-fertilized control.

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Figure 13. Biomass accumulation by hybrid bromegrass at Carrot River 1 (CR1) and

Carrot River 2 (CR2), and by timothy at Arborfield (ABR) and Choiceland (CHL) in 2013. Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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Nitrogen uptake and nitrogen use efficiency

Nitrogen uptake was monitored throughout the growing season (Fig. 14). Uptake of N by hybrid bromegrass (CR1 and CR2) apparently occurred early in the growing season such that crop N uptake from fall applied fertilizers was maximized by the first sampling date (14 d) and by day 28 for spring applied fertilizers at CR1. Data from CR1 were highly variable and the apparent reduction in N associated with the application of SuperU™

remains an unexplained anomaly. At CR2, small increases in total N uptake were observed following day 28 for both fall and spring applied fertilizers. It is possible that higher organic matter levels at CR2 as compared to CR1 (6.47 versus 3.92 %, respectively) may have contributed to an ongoing supply of N via organic matter mineralization, and hence ongoing N uptake. Uptake on N by timothy similarly was maximized early in the growing season with little difference between fall and spring applications at ABR. At CHL, trends suggest enhanced N uptake from spring versus fall fertilizer application.

Nitrogen use efficiency was calculated according to Havlin et al. (2005), where N uptake is determined by subtracting the N uptake from an unfertilized control from the N uptake from the fertilized crop to account for the contribution of N from mineralization of organic matter, as follows:

NUE = (Total N uptake from fertilized plot) − (N uptake from unfertilized plot) Rate of fertilizer applied

Nitrogen use efficiency values ranged from as low as 17.6 % to as high as 61.1 %,

but NUE responses were inconsistent and unpredictable between sites and treatments (Table 5). Failure to detect significant differences in NUE within sites likely reflects the relatively high variability in the data, as trends suggest higher NUE from spring-applied treatments. In general, hydrid bromegrass had higher NUE values for spring applied N treatments whereas NUE for timothy was similar for both fall and spring applied treatments.

Table 5. Nitrogen use efficiency for hybrid bromegrass and timothy grown at the four experimental sites in 2013.

Treatment Carrot River 1 Carrot River 2 Arborfield Choiceland Hybrid bromegrass Timothy

Nitrogen Use Efficiency (%)

Fall applied Urea 28.8 24.2 20.7 27.1

AGROTAIN® 38.0 26.2 26.7 28.9 SuperU™ 32.0 26.3 17.6 24.8 ALZON® 31.6 31.9 24.1 27.9

Spring applied Urea 52.3 61.1 27.7 25.1

AGROTAIN® 58.7 58.7 24.0 31.6 SuperU™ 21.9 46.8 31.6 33.4 ALZON® 43.3 47.2 18.7 37.8

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Figure 14. Impact of urease and nitrification inhibitors on uptake of N by hybrid

bromegrass at Carrot River 1 (CR1) and Carrot River 2 (CR2) and timothy at Arborfield (ABR) and Choiceland (CHL) following spring 2013 application of fertilizer treatments. Fall applied treatments received fertilizer in the fall of 2012, and were sampled at the same time as the spring applied treatments. Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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Seed yield

Fertilizer N application relative to the unfertilized control typically enhanced seed yields of both hybrid bromegrass and timothy; however, responses to specific EEF treatments were inconsistent between crops and sites (Fig. 15). Specifically, hybrid bromegrass was more responsive to fall-applied fertilizer than spring-applied fertilizer at both CR1 and CR2. The positive response of hybrid bromegrass to fall N fertilization is consistent of the reports of others (Loeppky and Coulman, 2001) and reflects the dual induction requirement for flowering that characterizes bromegrass, such that bromegrass sets reproductive tillers in the autumn for next years seed production (Heide, 1984). Thus, application of N in the fall presumably enhanced the potential for seed production in the following growing year, whereas response to spring fertilizer application was necessarily limited by the number of fertile tillers formed in the previous fall. In contrast, seed yield of timothy was unaffected by the timing of the fertilizer application. Timothy does not share the same dual induction requirement as hybrid bromegrass and seed yield potential is not set in the fall; thus timothy remains responsive to spring applied N.

Response to specific treatments varied between sites and crops. For example, the highest hybrid bromegrass seed yields (590 kg ha−1) were achieved with the fall application of the ALZON® at CR1, whereas none of the spring applied treatments, including ALZON®, differed significantly from the untreated control (139 kg ha−1). At CR2, the highest yield was achieved with the fall application of SuperU™, although differences between fall-applied treatments were not statistically significant. Spring application of SuperU™ at CR2 resulted in the lowest fertilized seed yields although only the seed yield associated with untreated spring applied urea was significantly different than all other spring applied fertilizers. Although trends suggest that timothy was responsive to some of the fertilizer treatments at ABR and CHL, treatment differences between fall-applied fertilizers or spring-applied fertilizers typically were not statistically significant.

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Figure 15. Seed yield response of hybrid bromegrass and timothy to fall and spring

application of untreated urea and enhanced efficiency N fertilizers. Treatments were as follows: C – unfertilized control; U – untreated urea; Su – SuperU™; Ag - AGROTAIN®; Al - ALZON®.

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Economic analyses of field experiments

Although few statistically significant seed yield responses were detected in the field experiments, an economic analyses using mean seed yields, was performed (Tables 6-9). We approached the economic analyses by considering whether or not the responses to the various EEF would be considered economically beneficial if each site and treatment represented the production experience from an individual farmer.

In hybrid bromegrass at both CR1 and CR2, the use of the EEF compared to untreated urea, applied in the fall, delivered positive returns in excess of the additional cost of the inhibitors. Spring application of EEF did not result in increased returns, with both SuperU™ and ALZON® resulting in a net loss relative to untreated urea at CR1 (Tables 6 and 7) and all three EEF resulting in a loss at CR2 when applied in the spring.

Table 6. Economic return achieved at Carrot River 1. Treatment Input Biomass Seed Revenue

($ ha-1) t ha-1 Return† kg ha-1 Return‡ Control 0 3.24 204.31 139 504.57 708.88 Fall applied

Urea 104.35 7.76 488.75 464 1684.32 2068.72 AGROTAIN® 121.09 8.73 549.74 556 2018.28 2446.93

SuperU™ 136.46 8.03 505.95 563 2043.69 2413.18 ALZON® 127.39 9.20 579.79 590 2141.70 2594.10

Spring applied Urea 104.35 9.17 577.46 292 1059.96 1533.07

AGROTAIN® 121.09 8.97 565.11 315 1143.45 1587.47 SuperU™ 136.46 7.98 502.61 254 922.02 1288.17 ALZON® 127.39 6.93 436.84 254 922.02 1231.47

† Return based on forage price of $63 tonne-1 (Hoimyr and Thompson, 2013) ‡ Return based on forage seed price of $3.63 kg-1 (Wong, 2013) Table 7. Economic return achieved at Carrot River 2. Treatment Input Biomass Seed Revenue


Urea 104.35 6.73 423.86 400 1452. 1771.51 AGROTAIN® 121.09 6.80 428.40 437 1586.31 1893.62

SuperU™ 136.46 7.02 442.01 496 1800.48 2106.03 ALZON® 127.39 6.89 433.94 443 1608.09 1914.64



† Return based on forage price of $63 tonne-1 (Hoimyr and Thompson, 2013) ‡ Return based on forage seed price of $3.63 kg-1 (Wong, 2013)

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Benefits associated with application of EEFs in timothy at Arborfield and Choiceland were inconsistent (Tables 8 and 9). At Arborfield, the most profitable application strategy was fall applied AGROTAIN®, providing additional revenue of $186 per ha over fall applied urea; all other fall and spring applied treatments produced similar or negative returns over untreated urea. At Choiceland, spring application of fertilizer provided to be the most profitable compared to fall application, and all spring-applied EEFs resulted in positive returns. Only AGROTAIN® resulted in positive returns over urea alone, when applied in the fall.

Table 8. Economic return achieved at Arborfield. Treatment Input Biomass Seed Revenue


Urea 104.35 4.64 292.19 580 1073.00 1260.84 AGROTAIN® 121.09 4.80 302.09 681 1259.85 1440.85

SuperU™ 136.46 4.84 304.79 592 1095.20 1263.53 ALZON® 127.39 3.96 249.17 564 1043.40 1165.18



† Return based on forage price of $63 tonne-1 (Hoimyr and Thompson, 2013) ‡ Return based on forage seed price of $3.63 kg-1 (Wong, 2013) Table 9. Economic return achieved at Choiceland. Treatment Input Biomass Seed Revenue


Urea 104.35 5.77 363.32 486 899.1 1158.07 AGROTAIN® 121.09 6.54 412.15 578 1069.3 1360.36

SuperU™ 136.46 6.06 382.03 493 912.05 1157.62 ALZON® 127.39 5.52 347.45 398 736.3 956.36



† Return based on forage price of $63 tonne-1 (Hoimyr and Thompson, 2013) ‡ Return based on forage seed price of $3.63 kg-1 (Wong, 2013)

Averaged over all sites, the added cost of the EEFs ($16.74 – $32.11 per ha) can be offset by an increase in seed yield of only 1–2%, which is below the statistical threshold (p < 0.05) used in this study. Indeed, when averaged across sites, the added profit

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associated with the use of the EEFs as compared to untreated urea in bromegrass seed production was $298 ha-1 (ranging from $141 ha-1 to $423 ha-1) for fall-applied products. In timothy seed production, however, the added profit associated with the use of the EEFs as compared to untreated urea was only $55 ha-1 (ranging from -$211 ha-1 to $330 ha-1) for spring-applied products. Thus, whereas our results suggest that there are potential benefits to be derived from the use of EEFs, particularly for fall applications, their effectiveness in forage seed production systems appears to be limited and somewhat unpredictable.

Controlled Environment Experiments

Ammonia Volatilization Chamber Experiments

Impact of soil pH on ammonia volatilization

The impact of soil pH on ammonia volatilization from various EEF products was examined by comparing existing variations in soil pH collected from the CR1 and CR2 field sites. The homogenized soils from CR1 and CR2 had pH values of 7.75 and 6.87, respectively. Soil also was collected from the ABR site, with a soil pH of 5.86. The ABR soil was limed with CaCO3 to a pH of 7.1. Untreated urea was compared to urea-based N sources amended with urease inhibitors (AGROTAIN® and Piazur®), a nitrification inhibitor (ALZON®), or both inhibitors (SuperU™).

In general, SuperU™ and Piazur® reduced NH3 emissions, with Alzon®, urea and AGROTAIN® having the highest emissions (Fig. 16). The pH difference in CR1 and CR2 soils, however, did not influence the magnitude of these losses, as emission patterns between both soils were very similar and generally low, with maximum losses equivalent to approximately 6% of the applied N. This was unexpected, because typically it is reported that ammonia volatilization losses increase with increasing soil pH (Sommer et al., 2004). Ammonia volatilization losses from the ABR soils were approximately three times higher than the losses from the Carrot River soils, and there was a more pronounced separation of SuperU® and Piazur® from other EEFs, with SuperU® associated with the lowest NH3 emissions. Higher NH3 emissions in the ABR soils was unexpected given the acidic conditions of the unlimed ABR soil (pH 5.86), as soil pH values of less than 7 are considered to limit NH3 emissions due to the transformation of NH3-N to NH4-N and subsequent adsorption to soil colloids (Sommer et al., 2004). Additionally, emissions from ABR soils under acidic conditions reached the emission peak 48 to 72 h earlier than the less acidic soils from CR1 and CR2 soils. These results indicate that soil pH is not necessarily a primary driver for governing NH3 emissions, and other factor(s) influence the magnitude of losses.

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Figure 16. Impact of soil pH on ammonia volatilization under controlled emissions chamber conditions.

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It has been suggested that soil urease activity may be an important factor governing ammonia volatilization (Rochette et al., 2009). Urease is a product of microbial activity and reportedly is enhanced in zero till soils (Rochette et al. 2009). We hypothesized that forage grass stands similarly may have high urease activity levels. Moreover, we suspected the soil urease activity to be one possible reason for the differences in NH3 emissions between CR1/CR2 and ABR soils, and tested our hypothesis by conducting a soil urease activity assessment according to Kandeler and Gerber (1988). The results of this assay revealed that urease activity was more than 10 times higher in the acidic ABR soil compared to the CR1/CR2-soils (data not shown), and this increase in urease activity was enhanced following liming and incubation of the ABR soil. On the other hand, there was no difference in urease activity between CR1 and CR2 soils. This analysis identified that soil urease activity was likely a more important factor controlling NH3 emissions than soil pH. Differences in urease activity between the ABR and CR1/CR2 soils are likely related to field management practices. Specifically, the forage stands at CR1/CR2 were burned annually in the spring, whereas the ABR site was not. Burning plant biomass in the field can significantly reduce the amount of organic matter that is available for decomposition, thereby reducing both microbial activity and the consequent amount of soil urease introduced to the soil.

Impact of soil water content on ammonia volatilization

Using the ABR soil, which had high ammonia emissions in the previous experiment, soil water content was varied to determine the impact of moisture on ammonia volatilization from the different fertilizer products (Figure 17). The highest emissions were observed when the soil was at 75% field capacity (FC), and declined at both higher (100% FC) and lower (50% FC) soil water contents. Irrespective of soil water content, SuperU™ and Piazur® were the most effective EEFs for reducing volatilization losses. It is interesting that Piazur® and AGROTAIN®, which both use a similar inhibitor, varied in effectiveness. Piazur® and AGROTAIN® differ in that AGROTAIN® is a surface applied product whereas Piazur® incorporates the inhibitor into the fertilizer granule during the manufacturing process.

Impact of soil temperature on ammonia volatilization

Using the ABR soil, soil temperature was manipulated to examine the impact of temperature on ammonia volatilization from the different EEF products (Figure 18). Emissions were enhanced as temperature was increased from 5oC to 26oC. Moreover, higher temperatures resulted in early ammonia emissions, with maximum emissions achieved within 72 h of fertilizer application. At 5oC, maximum emissions were delayed for approximately 250 h (i.e., more than 10 d). These observations are consistent with field experiment observations in which volatilization losses during the cool fall period typically were lower than ammonia volatilization losses experienced in the spring. Both SuperU™ and Piazur®, which contain a urease inhibitor, were most effective in reducing ammonia losses relative to the untreated urea control. Losses greater than 20% of the applied fertilizer were noted at 26oC.

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Figure 17. Impact of soil moisture content on ammonia volatilization under controlled emissions chamber conditions.

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Figure 18. Impact of soil temperature on ammonia volatilization under controlled

emissions chamber conditions.

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Assessing Gaseous N losses Using 15N labeled Urea

Ammonia emissions from 15N labeled urea were assessed over a 10-d sampling period under controlled conditions. Ammonia emissions from untreated urea, and urea treated with the nitrification inhibitor DCD increased rapidly following application (Fig. 19), peaking 4 d after fertilizer application. The inhibitor DCD inhibits the activity of nitrifying bacteria, and thus enhanced ammonia emissions likely occurred because the nitrification inhibitor effectively prevented the conversion of ammonium to nitrate, thereby maintaining or enhancing the size of the ammonium pool susceptible to volatilization losses. Both AGROTAIN®, and AGROTAIN® + DCD (a SuperU™ equivalent), successfully inhibited ammonia losses during the 10 day incubation time.

Figure 19. Impact of urease (AGROTAIN®) and nitrification inhibitors (DCD) on 15N

labeled ammonia fluxes under controlled environmental conditions.

Nitrous oxide fluxes were greatest from the untreated urea control (Fig. 20), peaking on day 7. AGROTAIN®, a urease inhibitor had higher flux values than the nitrification inhibitors; this result is not surprising as a urease inhibitor is not expected to have a direct effect on the N pool contributing to nitrous oxide emissions. Presumably the urease inhibitor slowed the conversion of urea to ammonia, but had no subsequent impact on the conversion of ammonia to nitrate.

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Figure 20. Impact of urease (AGROTAIN®) and nitrification (DCD) inhibitors on 15N

labeled nitrous oxide fluxes under controlled environmental conditions.

Emission factors for the 15NH3 captured during the 10-d incubation confirmed that both AGROTAIN®, and AGROTAIN® + DCD, reduced the proportion of 15N-labelled urea lost via ammonia volatilization (Fig. 21). The DCD alone increased ammonia emissions relative to the urease inhibitors, presumably by retaining more N in the susceptible ammonium pool. The emission factors for the N2O similarly suggested that the nitrification inhibitor DCD reduced losses via denitrification, although treatment differences were not statistically significant (Fig. 22). Overall, losses via ammonia volatilization were proportionally higher than losses associated with denitrification (Fig. 23), and thus treatments that included DCD were most successful in reducing overall emissions during the 10-d incubation period. Although differences in uptake by the plant were not statistically significant, those treatments that reduced gaseous emissions also appeared to promote enhanced 15N uptake in the plant tissue (Fig. 24).

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Figure 21. Emission factors for 15NH3 losses from 15N labeled urea and urea treated with

urease (AGROTAIN®) and nitrification (DCD) inhibitors under controlled environmental conditions.

Figure 22. Emission factors for 15N2O losses from 15N labeled urea and urea treated with


0

1

2

3

4

5

Ureaonly Urea+DCD Urea+Agrotain Urea+Agrotain+DCD

Percentofnettotalfertilizer15N

Fertilizerproduct

0.00

0.01

0.02

0.03

0.04

0.05


Percentofnettotalfertilizer15N

Fertilizerproduct

40

Figure 23. Percentage of total 15N labeled urea (i.e., 15NH3 + 15N2O) lost via gaseous

emissions from 15N labeled urea and urea treated with urease (AGROTAIN®) and nitrification (DCD) inhibitors under controlled environmental conditions.

Figure 24. Emission factors for 15NH3 losses from 15N labeled urea and urea treated with


0

1

2

3

4

5


%15Nlostthroughemissions

Fertilizerproduct

emissionfactor15NH3%

emissionfactor15N2O%

0

5

10

15

20

25

30

35

40


Percentofnettotal1

5Napplied

Fertilizerproduct

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9. CONCLUSIONS AND RECOMMENDATIONS

Field and laboratory studies were conducted to examine the potential agronomic and environmental benefits of enhanced efficiency fertilizers (EEFs; employing urease and nitrification inhibitors, alone and in combination) in forage seed production in the Parkland region of Saskatchewan. Field sites were established in existing forage seed fields at four locations (Carrot River 1, Carrot River 2, Arborfield and Choiceland) in the Parkland region of Saskatchewan to evaluate the impact of fall and spring applications of urea and three enhanced efficiency fertilizers urea fertilizers (EEFs) on NH3 volatilization, N2O emissions, nitrogen use efficiency (NUE), and consequent seed yield of hybrid bromegrass and timothy. Additionally, a series of laboratory incubation and growth chamber experiments further investigated factors that influence the success of EEFs in reducing gaseous N losses.

Field experiments revealed the efficacy of EEF N products in established hybrid bromegrass stands varied depending on whether the product was applied in the fall or spring. Fall application of the EEFs were generally more beneficial, particularly in hybrid bromegrass stands—increasing dry matter and seed yields by 4 to 11% and 15 to 22%, respectively, compared to urea alone. Conversely, spring applications generally resulted in decreased dry matter (3–12%) and seed (2–22%) yields. These results were not unexpected, as bromegrass forms reproductive tillers in the fall. As well, vernalization of the tillers is required to induce reproductive growth. Thus, fall fertilizer applications are timed to enhance tillering, which in turn, increases flowering and seed production. Use of the EEFs appeared to delay N-transformations that could lead to N losses and potentially reduce the size of the plant-available N pool. Indeed, use of the EEFs resulted in small increases in NUE (2–5%) compared to the urea. In general, however, differences between urea and the EEFs were not statistically significant (p > 0.05).

Timothy production was greatest when fertilizer (urea or EEFs) was applied in the spring, though in general, the effects of the EEF products were not significant (p > 0.05). Interestingly, however, the effects of the EEFs on timothy production appeared to be site-specific, with a general negative (12–23%) effect at Arborfield and a positive (10–30%) effect at Choiceland.

From an economic standpoint, the added cost of the EEFs ($16.74 – $32.11 per ha) can be offset by an increase in seed yield of only 1–2%, which is below the statistical threshold (p < 0.05) used in this study. Indeed, when averaged across sites, the added profit associated with the use of the EEFs in bromegrass seed production was $298 ha-1 (ranging from $141 ha-1 to $423 ha-1) for fall-applied products. In timothy seed production, however, the added profit associated with the use of the EEFs was only $55 ha-1 (ranging from -$211 ha-1 to $330 ha-1) for spring-applied products. Thus, whereas our results suggest that there are potential benefits to be derived from the use of EEFs, their effectiveness appears to be both site specific and dependent on environmental conditions.

Laboratory and growth chamber studies revealed that urease activity associated with organic matter accumulation is an important factor controlling the potential for ammonia

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volatilization losses. Although high soil pH is frequently cited as a main factor controlling ammonia volatilization losses, laboratory incubation experiments revealed that high volatilization losses can occur even on acid soils if high levels of urease are present. Thus, in existing grass forage stands where grass residues are retained on the surface (i.e., are not burned in the early spring) high urease activity is likely to promote ammonia volatilization losses. Under these circumstances, application of an EEF containing a urease inhibitor may be warranted.

10. SUCCESS STORIES/PRACTICAL IMPLICATIONS FOR PRODUCERS OR INDUSTRY.

This project contributed to an improved understanding of N management in forage grass seed production to enhance N use efficiency and promote enhanced seed yields while minimizing environmental impacts. Importantly, although not conclusive, results suggest that some benefits may be gained from using EEFs containing nitrification or urease inhibitors, particularly when conditions are optimal for N losses either via denitrification or ammonia volatilization. 11. PATENTS/IP GENERATED/COMMERCIALIZED PRODUCTS

No patents or commercialized products were developed. Information for public dissemination was developed. 12. TECHNOLOGY TRANSFER ACTIVITIES Technology transfer has occurred through ongoing meetings with farmers (field days and annual workshop) and industry representatives. Specific activities were as follows: Farrell, R.E. 2014. Reducing Greenhouse Gas Emissions through 4R Nutrient Stewardship: A Saskatchewan Perspective. International Stewardship Symposium: Feeding Crops to Feed the World, 14–16 July, Saskatoon, SK (Invited presentation). Farrell, R.E. 2014. International Stewardship Symposium – Pre-Conference field research tour. “Local examples of research on the ground utilizing 4R Nutrient Stewardship”. July 14th (46 attendees). Walley, F.L. 2014. Improving yields in forage stands by enhancing nitrogen use efficiency. Presented at the Saskatchewan Ministry of Agriculture Regional Extension Staff Update; November 19, 2014 (approx. 20 SMA staff in attendance). Walley, F.L. 2014. Nitrogen cycling in soils. Presented at the Agronomy Research Update; December 11 and 12, 2014 (approx. 200 agrologists in attendance). Yannikos, N., F.L. Walley and R.E. Farrell. 2014. Assessing the Impact of Stabilized Urea Fertilizers on Ammonia Volatilization Under Controlled Environment and Soil Conditions. ASA, CSSA, & SSSA 2014 International Annual Meetings, 2–5 Nov., Long Beach, CA (Oral presentation).

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Walley, F.L. 2014. Improving yields by enhancing nitrogen use efficiency. Presented at the 2014 MFSA Forage Seed Conference, sponsored by the Manitoba Forage Seed Association; January 13, 2014 (approx. 150 producers in attendance). Walley, F.L. 2014. Presented “Effects of improved nitrogen and stand management on net GHG balances in forage productions systems” at the Agriculture and Agri-Food Canada (AAFC) / Agriculture Greenhouse Gas Program Grasslands (AGGP) Network Meeting Report, Saskatoon Research Centre, on March 20, 2014. Attendees discussed ongoing research, and identified gaps, including BMPs. Yannikos, N. J. Woodhouse, F. Walley and R. Farrell. 2013. Assessment of urease and nitrification inhibitors for improving nitrogen use efficiency and yield in forage seed production: Project update. Saskatchewan Forage Seed Development Commission Forage Seed Information Workshop and SFSDC AGM. Dec. 5, Nipawin, SK. Woodhouse, J., F.L. Walley, and R.E. Farrell. 2013. Synchronizing nitrogen application with uptake using urease and nitrification inhibitors to maximize nitrogen use in forage seed stands in Northeastern Saskatchewan. Oral presentation at the 2013 ASA-CSSA-SSSA International Annual Meetings, Nov. 3–6, Tampa Bay, FL. Yannikos, N., Fran L. Walley and Richard Farrell. 2013. Assessing the efficacy of nitrification and urease inhibitors on nitrous oxide and ammonia emissions in forage seed production in the Saskatchewan Parkland region. Poster presentation at the 2013 ASA-CSSA-SSSA International Annual Meetings, Nov. 3–6, Tampa Bay, FL. Yannikos, Nils and James Woodhouse. 2013. Presented an overview of the project during the Forage Seed Research Field Day at Melfort, July 24, 2013 (approximately 200 participants in attendance). Farrell, R.E. 2013. Use of nitrification and urease inhibitors in forage seed production. Presented at the 2013 Agronomy Research Update, sponsored by the Saskatchewan Ministry of Agriculture, Prairie Certified Crop Advisor Board and the University of Saskatchewan; December 11, 2013 (70 extension agents in attendance). Farrell, R.E., N. Yannikos (Ph.D. candidate) and James Woodhouse (M.Sc. candidate) attended and participated in a special AGGP session at the 2013 Canadian Society of Soil Science Annual Meeting, Winnipeg, MB, July 22-25. Farrell co-chaired the session. All three also participated in a related meeting of AGGP participants to discuss linkages, and identify gaps and potential future directions. Dr. Farrell demonstrated the new FTIR-mga system for automated GHG measurements in the field during a field tour conducted for members of the Global Research Alliance. June 8, 2012.

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Walley, F. 2012. Presentation on “Nitrogen Use Research in Forage Seed Crops” at the Saskatchewan Forage Seed Development Commission (SFSDC) Annual General Meeting and Information Session was held in Nipawin, SK on December 5, 2012. Walley, F. 2012. Improving nitrogen use efficiency in forage seed production. Saskatchewan Forage Seed Development Commission, Prairie Seeds Newsletter, November 2012, p. 6. Farrell, R.E. and F.L. Walley. 2012. Presentation on use of nitrification and urease inhibitors in forage seed production, Forage Seed Field Day (Nipawin), Sponsored by the Saskatchewan Forage Seed Development Commission, July 25, 2012. Walley, F. and R. Farrell. 2011. Use of nitrification and urease inhibitors in forage seed production: Project update. Saskatchewan Forage Seed Development Commission Forage Seed Information Workshop and SFSDC AGM, Evergreen Centre, Nipawin, SK, December 6, 2011 (20 farmers in attendance). 13. INDUSTRY CONTRIBUTIONS OR SUPPORT RECEIVED

The Saskatchewan Forage Seed Growers Association provided $4,000 cash towards this project. 14. IS THERE A NEED TO CONDUCT FOLLOW-UP RESEARCH?

Results of this study indicate that the efficacy of EEFs products in established hybrid bromegrass stands varied depending on whether the product was applied in the fall or spring, and depending on a myriad of site-specific soil and environmental characteristics. Nonetheless, data suggest that under some conditions, EEFs may effectively reduce gaseous losses from N fertilizer. Reducing gaseous N losses has both environmental and agronomic implications and thus further research regarding the effective use of EEFs in various cropping systems is warranted. 15. ACKNOWLEDGEMENTS

The support from the Agriculture Development Fund, Saskatchewan Ministry of Agriculture was acknowledged in all presentations listed above. Additionally, we acknowledge the support and contributions from the following:

The cooperation of forage growers Marcel Enns, Bernie Schultz, and Kyle Schmidt is gratefully acknowledged.

The assistance and advice provided by Clayton Myhre, Pickseed Canada, is gratefully acknowledged.

Financial Support from the Saskatchewan Forage Seed Development Commission is gratefully acknowledged.

16. APPENDICES

None

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